Organic compounds

ABSTRACT

A compound of formula (I) 
     
       
         
         
             
             
         
       
     
     wherein
 
n is 0 to 4;
 
R is ethyl or vinyl;
 
R 1  is hydrogen or (C 1-6 )alkyl,
 
R 2  is hydrogen or
         cycloalkyl including (C 3-6 )cycloalkyl, or   unsubstituted (C 1-6 )alkyl, or   (C 1-6 )alkyl substituted by one or more of
           hydroxy; preferably one or two,   halogen,   (C 3-6 )cycloalkyl, or
 
R 1  is hydroxy and R 2  is formyl.

CROSS REFERENCE TO RELATED APPLICATIONS

This applications claims to benefit and priority to European patentapplication 07450053.9, filed on Mar. 20, 2007, which is herebyincorporated by reference in its entirety.

The present invention relates to organic compounds, namelypleuromutilins.

Pleuromutilin, a compound of formula A

is a naturally occurring antibiotic, e.g. produced by the basidomycetesPleurotus mutilus and P. passeckerianus, see e.g. The Merck Index, 13thedition, item 7617. A number of further pleuromutilins having theprinciple ring structure of pleuromutilin and being substituted at thehydroxy group have been developed, e.g. as antimicrobials.

From WO 02/04414 A1 pleuromutilin derivatives, e.g.14-O-[(Aminocyclohexan-2-yl (and -3-yl)-sulfanyl)-acetyl]-mutilins; fromWO 07/014409 A1 e.g. 14-O-[((Mono- ordialkylamino)-cycloalkylsulfanyl)-acetyl]-mutilins and from WO 07/000004A1 e.g. [((Acyl-hydroxy-amino)cycloalkylsulfanyl)-acetyl]-mutilins, areknown.

We have now found pleuromutilins with interesting activity combined withan unexpexted remarkable metabolic stability.

The pleuromutilin derivatives according to the invention are compoundsof formula (I)

whereinn is 0 to 4;R is ethyl or vinyl;R₁ is hydrogen or (C₁₋₆)alkyl,R₂ is hydrogen or

-   -   cycloalkyl including (C₃₋₆)cycloalkyl, or    -   unsubstituted (C₁₋₆)alkyl, or    -   (C₁₋₆)alkyl substituted by one or more of        -   hydroxy; preferably one or two,        -   halogen,        -   (C₃₋₆)cycloalkyl, or            R₁ is hydroxy and R₂ is formyl.

Preferred compounds of the present invention are

-   -   a compound of formula (II)

-   -   a compound of formula (III)

and

-   -   a compound of formula (IV)

wherein n, R₁ and R₂ are as defined above.

Particularly preferred is a compound selected from the group consistingof

14-O-{[(1R, 2R, 4R)4-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinand the (1S, 2S, 4S) diastereomer thereof

14-O-{[(1R, 2R, 5S)-5-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5R) diastereomer thereof

14-O-{[(1R, 2R,4S)-4-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the (1S,2S, 4R) diastereomer thereof

14-O-{[(1R, 2R,5R)-5-Amino-2-hydroxy-cyclobexylsulfanyl]-acetyl}-mutilin and the (1S,2S, 5S) diastereomer thereof

14-O-{[(1R, 2R,3R)-3-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the (1S,2S, 3S) diastereomer thereof

14-O-{[(1R, 2R,4R)-4-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the(1S, 2S, 4S) diastereomer thereof

14-O-{[(1R, 2R,4R)4-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the(1S, 2S, 4S) diastereomer thereof

14-O-{[(1R, 2R,5S)-5-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the(1S, 2S, 5R) diastereomer thereof

14-O-{[(1R, 2R,5S)-5-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the(1S, 2S, 5R) diastereomer thereof

14-O-{[(1R, 2R,4S)-4-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the(1S, 2S, 4R) diastereomer thereof

14-O-{[(1R, 2R,5R)-5-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the(1S, 2S, 5S) diastereomer thereof

14-O-{[(1R, 2R,3R)-3-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the(1S, 2S, 3S) diastereomer thereof

14-O-{[(1R, 2R,3R)-3-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the(1S, 2S, 3S) diastereomer thereof

14-O-{[(1R, 2R,4S)4-(Formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinand the (1S, 2S, 4R) diastereomer thereof

14-O-{[(1R, 2R,5S)-5-(Formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinand the (1S, 2S, 5R) diastereomer thereof

and

14-O-{[(1R, 2R,3R/S)-3-(Formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinand the (1S, 2S, 3R/S) diastereomer thereof.

A compound provided by the present invention is herein also designatedas “compound(s) of (according to) the present invention”. A compound ofthe present invention includes a compound in any form, e.g. in freeform, in the form of a salt, in the form of a solvate and in the form ofa salt and a solvate.

According to another aspect, the present invention provides a compoundof the present invention in the form of a salt and/or solvate.

The salts include preferably pharmaceutically acceptable salts, althoughpharmaceutically unacceptable salts are included, e.g. forpreparation/isolation/purification purposes.

A salt of a compound of the present invention includes a base salt or anacid addition salt. Pharmaceutically acceptable base salts includeammonium salts such as trimethylammonium salt, alkali metal salts suchas those of sodium and potassium, alkaline earth metal salts such asthose of calcium and magnesium, and salts with organic bases, includingsalts of primary, secondary and tertiary amines, such as isopropylamine,diethylamine, ethanolamine, trimethylamine, dicyclohexyl amine andN-methyl-D-glucamine, preferably sodium salts. Acid addition saltsinclude salts of a compound of the present invention with an acid, e.g.hydrogen fumaric acid, fumaric acid, tartaric acid,ethane-1,2-disulphonic acid, maleic acid, naphthalin-1,5-sulphonic acid,acetic acid, maleic acid, suceinic acid, salicylic acid, azelaic acid,2-[(2,6-dichlorophenyl)amino]bezene acetic acid, hydrochloric acid,deuterochloric acid, preferably hydrochloric acid.

A compound of the present invention in free form may be converted into acorresponding compound in the form of a salt, and vice versa. A compoundof the present invention in free form or in the form of a salt and/or inthe form of a solvate may be converted into a corresponding compound infree form or in the form of a salt in non-solvated form, and vice versa.

A compound of the present invention may exist in the form of isomers andmixtures thereof, e.g. optical isomers, diastereoisomers, cis/transconformers. A compound of the present invention may e.g. containasymmetric carbon atoms and may thus exist in the form of enatiomers ordiastereoisomers and mixtures thereof, e.g. racemates or diastereomericmixtures. Any asymmetric carbon atom may be present in the R)-, (S)- or(R,S)-configuration, preferably in the (R)- or (S)-configuration.

For example, in a compound of formula I the carbon atom of thecycloalkyl ring which is attached to the sulphur atom, the carbon atomof the cycloalkyl ring which is attached to the hydroxy group, and thecarbon atom of the cycloalkyl ring to which the (CH₂)_(n)N(R₁R₂) groupis attached, all are asymmetric carbon atoms. Substituents attached tosuch asymmetric carbon atom may thus exist in (R) and (S) configuration,including mixtures thereof. For example, if in a compound of formula IR₂ is substituted alkyl and that substituent is attached to a carbonatom of the side chain of such alkyl, the carbon atom to which suchsubstituent is attached is an asymmetric carbon atom and suchsubstituent may be in the (R)- and (S)-configuration, including mixturesthereof.

The configuration of substituents attached to asymmetric carbon atoms ofthe mutilin-ring is preferably the same as in natural pleuromutilin.

Isomeric mixtures may be separated as appropriate, e.g. according, e.g.analogously, to a method as conventional, to obtain pure isomers. Thepresent invention includes a compound of the present invention in anyisomeric form and in any isomeric mixture. The present invention alsoincludes tautomers of a compound of the present invention, wheretautomers can exist.

The compounds of the present invention exhibit pharmacological activityand are therefore useful as pharmaceuticals.

For example, the compounds of the present invention show antimicrobial,e.g. antibacterial, activity against gram positive bacteria, such ascoagulase positive Staphylococci, e.g. Staphylococcus aureus, coagulasenegative Staphylococci, e.g. Staphylococcus epidermidis, Staphylococcushaemolyticus, and Streptococci, e.g. Streptococcus pyogenes,Streptococcus pneumoniae, Enterococci, e.g. Enterococcus faecium andListeria monocytogenes and against gram negative bacteria such asMoraxella, e.g. Moraxella catarrhalis, and Haemophilus, e.g. Haemophilusinfluenzae, and Legionella, e.g. Legionella pneumophila, Neisseriaceae,e.g. Neisseria gonorrhoeae, as well as against Mycoplasms, Chlamydia andobligatory anaerobes, e.g Bacteroides fragilis, Clostridium difficile,Fusobacterium spp., and Propionibacterium spp.

The in vitro activity against aerobic bacteria was determined by AgarDilution Test or Microdilution Test according to the Clinical andLaboratory Standards Institute (CLSI, former NCCLS) Document M7-A7 Vol.26, No. 2: “Methods for dilution Antimicrobial Susceptibility Tests forBacteria that Grow Aerobically—Approved Standard; Seventh Edition(2006)”; and the test against anaerobic bacteria was performed accordingto the Clinical and Laboratory Standards Institute (CLSI, former NCCLS),Document, M11-A≢, Vol. 24, No. 2: “Methods for AntimicrobialSusceptibility Testing of Anaerobic Bacteria—Approved Standard; SixthEdition (2004)” and the in vivo activity was tested by the septicaemiamouse model against Staphylococcus aureus.

Compounds of the present invention are therefore suitable for thetreatment and prevention of diseases which are mediated by microbes,e.g. by bacteria. Diseases which may also be treated include e.g.diseases mediated by Helicobacter, such as Helicobacter pylori, anddiseases mediated by Mycobacterium tuberculosis. Diseases which may alsobe treated include in general inflammatory diseases, where microbes aremediating said inflammation, e.g. including acne.

In another aspect the present invention provides a compound of thepresent invention for use as a pharmaceutical, preferably as anantimicrobial, such as an antibiotic, e.g. and an anti-anaerobic.

In another aspect the present invention provides a compound of thepresent invention for use in acne treatment.

In a further aspect the present invention provides a compound of thepresent invention for use in the preparation of a medicament for thetreatment of diseases, mediated by microbes, such as bacterials, forexample

diseases mediated by bacteria, e.g. selected from Staphylococci,Streptococci, Enterococci;

diseases mediated by bacteria, e.g. selected from Moraxella,Haemophilus, Legionella, Neisseriaceae;

diseases mediated by Helicobacter;

diseases mediated by Mycobacterium tuberculosis;

e.g. diseases mediated by Mycoplasms, Chlamydia and obligatoryanaerobes;

and for the treatment of acne.

In a further aspect the present invention provides a method of treatmentof diseases mediated by microbes which comprises administering to asubject in need of such treatment an effective amount of a compound ofthe present invention e.g. in the form of a pharmaceutical composition.

In a further aspect the present invention provides a method of treatmentof acne which comprises administering to a subject in need of suchtreatment an effective amount of a compound of the present inventione.g. in the form of a pharmaceutical composition.

Treatment includes treatment and prophylaxis.

For antimicrobial and acne treatment, the appropriate dosage will, ofcourse, vary depending upon, for example, the chemical nature and thepharmakokinetic data of a compound of the present invention employed,the individual host, the mode of administration and the nature andseverity of the conditions being treated. However, in general, forsatisfactory results in larger mammals, for example humans, an indicateddaily dosage is in the range from about 0.5 mg to 3 g of a compound ofthe present invention conveniently administered, for example, in divideddoses up to four times a day.

A compound of the present invention may be administered by anyconventional route, for example enterally, e.g. including nasal, buccal,rectal, oral administration; parenterally, e.g. including intravenous,intramuscular, subcutaneous administration; or topically, e.g. includingepicutaneous, intranasal, intratracheal administration, e.g. in form ofcoated or uncoated tablets, capsules, injectable solutions orsuspensions, e.g. in the form of ampoules, vials, in the form of creams,gels, pastes, inhaler powder, foams, tinctures, lip sticks, drops,sprays, or in the form of suppositories, e.g. in analogous manner tomacrolides, such as erythromycins, e.g. clarithromycin or azithromycin.

A compound of the present invention may be administered in the form of apharmaceutically acceptable salt, e.g. an acid addition salt or a baseaddition salt, e.g. a metal salt, or in free form, optionally in theform of a solvate. A compound of the present invention in the form of asalt exhibits the same order of activity as the compound in free form,optionally in the form of a solvate.

A compound of the present invention may be used for pharmaceuticaltreatment according to the present invention alone or in combinationwith one or more other pharmaceutically active agents. Such otherpharmaceutically active agents include e.g. other antibiotics andantiinflammatory agents, and, if a compound of the present invention isused in the treatment of acne, other pharmaceutically agents includefurthermore agents which are active against acne.

Combinations include fixed combinations, in which two or morepharmaceutically active agents are in the same formulation; kits, inwhich two or more pharmaceutically active agents in separateformulations are sold in the same package, e.g. with instruction forco-administration; and free combinations in which the pharmaceuticallyactive agents are packaged separately, but instruction for simultaneousor sequential administration are given.

In another aspect the present invention provides a pharmaceuticalcomposition comprising a compound of the present invention in free formor in the form of a pharmaceutically acceptable salt and/or in the formof a solvate in association with at least one pharmaceutical, excipient,e.g. carrier or diluent, e.g. including fillers, binders,disintegrators, flow conditioners, lubricants, sugars and sweeteners,fragrances, preservatives, stabilizers, wetting agents and/oremulsifiers, solubilizers, salts for regulating osmotic pressure and/orbuffers.

In another aspect the present invention provides a pharmaceuticalcomposition according to the present invention, further comprisinganother pharmaceutically active agent.

Such pharmaceutical compositions may be manufactured according, e.g.analogously, to a method as conventional, e.g. by mixing, granulating,coating, dissolving or lyophilizing processes. Unit dosage form maycontain, for example, from about 0.5 mg to about 2000 mg, such as 10 mgto about 500 mg.

The compounds of the present invention are additionally suitable asveterinary agents, e.g. veterinary active compounds, e.g. in theprophylaxis and in the treatment of microbial, e.g. bacterial diseases,in animals, such as fowl, pigs and calves, e.g., and for diluting fluidsfor artificial insemination and for egg-dipping techniques.

In another aspect the present invention provides a compound of thepresent invention for use as a veterinary agent.

In a further aspect the present invention provides a compound of thepresent invention for the preparation of a veterinary composition whichis useful as a veterinary agent.

In another aspect the present invention provides a veterinary method forthe prophylaxis and the treatment of microbial, e.g. bacterial diseaseswhich comprises administering to a subject in need of such treatment aneffective amount of a compound of the present invention, e.g. in theform of a veterinary composition.

Examples 1 to 15 following thereafter exhibit MICs≦2 μg/ml againstStaphylococcus aureus ATCC49951 and Streptococcus pneumoniae ATCC49619.

The metabolic stability for compounds of the present invention wasdetermined by using cryopreserved primary human hepatocytes. 1×10⁶cells/mL were incubated in the absence and the presence of 5 and 25μg/mL of the test compounds at 37° C., 5% CO₂ for 4 hours, To evaluatethe in vitro degradation under assay conditions, a sample of each testcompound was incubated also in the absence of hepatocytes. Theincubation was stopped by freezing the reaction mixture. Afterultrafiltration and washing of the filter with acetonitrile, the samplesolution was analyzed for parent compound disappearance or metaboliteappearance using LC/MS (ion trap). The metabolic stability valuecorresponds to the detected parent compound in % after incubation.

At the compounds of the present invention, the introduction of a hydroxygroup in ortho position to the sulphur substituent in the cyclohexylring reveals unexpected improvements in metabolic stability of themicrobiologically active components. Parent compound or activemetabolite were more stable after incubation with primary humanhepatocytes in comparison to derivatives without a hydroxy group in thecyclohexyl moiety of the pleuromutilin side chain.

For example after 4 h incubation with human hepatocytes at a compoundconcentration of 5 μg/mL, for a mixture of 14-O-{[(1R, 2R,5S)-5-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin hydrochlorideand the (1S, 2S, 5R) diastereomer hydrochloride thereof—Example 2 of thepresent invention—66% of parent compounds were found, whereas formixture of 14-O-{[(1R, 3S)-3-Amino-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride and the (1S, S3R) diastereomer hydrochloridethereof—analogous derivative without hydroxyl group—only 24% of parentcompounds could be detected.

EXAMPLES

The trivial name mutilin refers to the IUPAC systematic name (1S, 2R,3S, 4S, 6R, 7R, 8R,14R)-3,6-dihydroxy-2,4,7,14-tetramethyl-4-vinyl-tricyclo[5.4.3.0^(1,8)]tetradecan-9-one.In the examples, pleuromutilin derivatives are numbered in analogy tothe mutilin numbering system described by H. Berner (Berner, H.; Schulz,G.; Schneider H. Tetrahedron 1980, 36, 1807-1811.):

Pleuromutilin thiol and pleuromutilin tosylate are compounds offormulae:

Example 1 14-O-{[(1R, 2R,4R)4-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 4S) diastereomer hydrochloride

Step A1. 14-O-{[(1R, 2R,4R)-4-tert-Butoxycarbonylamino-2-hydroxy-cyclohexylsuffanyl]-acetyl}-mutilin+(1S,2S, 4S) diastereomer and 14-O-{[(1R, 2R,5S)-5-tert-Butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer and

14-O-{[(1R, 2R,4S)-4-tert-Butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer

To a solution of 3,4-epoxycyclohexyl-carbamic acid tert-butyl ester(Gómez-Sánchez, E.; Marco-Contelles J. Tetrahedron 2005, 61, 1207-1219.)(4.27 g, 20 mmol) and pleuromutilin thiol (Nagarajan, R. Eli Lilly andCompany 1978, U.S. Pat. No. 4,130,709) (7.10 g, 18 mmol) in 200 ml oftetrahydrofuran was added aluminum oxide (40 g, Brockmann activity I,neutral) and the resulting mixture was stirred for 40 hours at roomtemperature. The suspension was filtered and concentrated under reducedpressure. The residue was subjected to chromatography (silica,cyclohexane/ethyl acetate=1/1) to yield 14-O-{[(1R, 2R,4R)-4-tert-butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4S) diastereomer (a) (R_(f)=0.38, 1.34 g, 12%) as well as a mixtureof 14-O-{[(1R, 2R,5S)-5-tert-butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer and 14-O-{[(1R, 2R,4S)-4-tert-butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer (b) (R_(f)=0.26, 2.81 g, 25%) as colorlessamorphous foams.

(a): ¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.74 (d, 1H, NH, J=7Hz), 6.13 (dd, 1H, 19-H, J=11 Hz and 18 Hz), 5.54 (d, 1H, 14-H, J=8 Hz),5.05 (m, 2H, 20-H), 4.90 (d, 1H, 2′-OH, J=5 Hz), 4.48 (d, 1H, 11-OH, J=6Hz), 3.55-3.20 (m, 6H, 1′-H, 2′-H, 4′-H, 11-H, 22-H), 2.40 (bs, 1H,4-H), 1.36 (s, 3H, 15-CH₃), 1.35 (s, 9H, tert-butyl), 1.06 (s, 3H,18-CH₃), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.62 (d, 3H, 16-CH₃, J=7 Hz).MS-ESI (m/z): 630 (MNa⁺), 1237 (2MNa⁺).

(b): ¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.70 (d, 1H, NH, J=7Hz), 6.12 (dd, 1H, 19-H, J=11 Hz and 18 Hz), 5.34 (d, 1H, 14-H, J=8 Hz),5.05 (m, 2H, 20-H), 4.82, 4.78 (d, 1H, 2′-OH, J=4 Hz), 4.48 (d, 1H,11-OH, J=6 Hz), 3.55-3.20 (m, 5H, 1′-H, 2′-H, 4′-H, 11-H, 22-H), 2.97(m, 1H, 1′-H), 2.40 (bs, 1H, 4-H), 1.35 (s, 12H, 15-CH₃, tert-butyl),1.05 (s, 3H, 18-CH₃), 0.82 (d, 3H, 17-CH₃, J=7 Hz), 0.62 (d, 3H, 16-CH₃,J=7 Hz). MS-ESI (m/z): 630 (MNa⁺), 1237 (2MNa⁺).

or Step A2. 14-O-{[(1R, 2R,4R)-4-tert-Butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4S) diastereomer and

14-O-{[(1R, 2R,5S)-5-tert-Butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer and14-O-{[(1R, 2R,4S)-4-tert-Butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer

To a solution of 3,4-epoxycyclohexyl-carbamic acid tert-butyl ester (10g, 47 mmol) and pleuromutilin thiol (16.6 g, 42 mmol) in 200 ml ofmethanol and 20 ml of dioxane was added 2N NaOH (21 ml, 42 mmol) and theresulting mixture was stirred for 16 hours at room temperature. Aftercompletion of the reaction the pH was set to 7 with diluted HCl and thereaction mixture was concentrated under reduced pressure. The residuewas diluted with water and brine and extracted three times with ethylacetate. The organic layers were dried over sodium sulfate and filtered.The filtrate was concentrated under reduced pressure and afterchromatography (silica, cyclohexane/ethyl acetate=1/1) 14-O-{[(1R, 2R,4R)-4-tert-butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4S) diastereomer (R_(f)=0.40, 3.1 g, 12% yield) as well as a mixtureof 14-O-{[(1R, 2R,5S)-5-tert-butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer and 14-O-{[(1R, 2R,4S)-4-tert-butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer (R_(f)=0.25, 6.35 g, 25%) were obtained ascolorless amorphous foams.

Step B. 14-O-{[(1R, 2R,4R)4-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 4S)diastereomer

To a solution of 14-O-{[(1R, 2R,4R)-4-tert-butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4S) diastereomer (1.34 g, 2.20 mmol) in 75 ml of dichloromethane wasadded trifluoroacetic acid (4 ml) at 4° C. and stirred for 5 hours atroom temperature. The reaction mixture was diluted with dichloromethaneand cautiously poured into a saturated NaHCO₃ solution. The phases wereseparated and the aqueous layer was washed two times withdichloromethane. The combined organic layers are dried over sodiumsulfate and filtered. After chromatography (silica, ethylacetate/methanol/35% ammonia solution=50/50/1) 14-O-{[(1R, 2R,4R)-4-amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 4S)diastereomer (745 mg, 67% yield) was obtained as colorless amorphousfoam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.14 (dd, 1H, 19-H, J=11Hz and 18 Hz), 5.54 (d, 1H, 14-H, J=8 Hz), 5.05 (m, 2H, 20-H), 4.50 (d,1H, 11-OH, J=6 Hz), 3.50-3.20 (m, 5H, 2′-H, 4′-H, 11-H, 22-H), 2.55 (m,1H, 1′-H), 2.40 (bs, 1H, 4-H), 1.35 (s, 3H, 15-CH₃), 1.06 (s, 3H,18-CH₃), 0.82 (d, 3H, 17-CH₃, J=7 Hz), 0.62 (d, 3H, 16-CH₃, J=7 Hz).MS-ESI (m/z): 508 (MH⁺), 530 (MNa⁺), 1015 (2MH⁺), 1037 (2MNa⁺).

Step C. 14-O-{[(1R, 2R,4R)-4-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 4S) diastereomer hydrochloride

A solution of 14-O-{[(1R, 2R,4R)-4-amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 4S)diastereomer (325 mg, 0.64 mmol) in 20 ml of dioxane was treated with 1NHCl (0.64 ml, 0.64 mmol). After stirring at room temperature for 30minutes the solution was lyophilized to obtain 14-O-{[(1R, 2R,4R)-4-amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 4S) diastereomer hydrochloride (quantitativeyield) as colorless amorphous solid.

¹H NMR (500 MHz, DMSO-d₆, δ, ppm, inter alia): 7.6 (bs, 3H, NH₃ ⁺), 6.14(dd, 1H, 19-H, J=11 Hz and 18 Hz), 5.55 (d, 1H, 14-H, J=8 Hz), 5.05 (m,2H, 20-H), 4.52 (d, 1H, 11-OH, J=6 Hz), 3.50-3.20 (m, 4H, 2′-H, 11-H,22-H), 3.03 (m, 1H, 4′-H), 2.53 (m, 1H, 1′-H), 2.40 (bs, 1H, 4-H), 1.37(s, 3H, 15-CH₃), 1.06 (s, 3H, 18-CH₃), 0.82 (d, 3H, 17-CH₃, J=7 Hz),0.62 (d, 3H, 16-CH₃, J=7 Hz). MS-ESI (m/z): 508 (MH⁺), 530 (MNa⁺), 1015(2MH⁺), 1037 (2MNa⁺), 542 (MCl⁻).

Example 2 14-O-{[(1R, 2R,5S)-5-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1s, 2S, 5R) diastereomer hydrochloride

Step A. 14-O-{[(1R, 2R,5S)-5-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 5R)diastereomer and

14-O-{[(1R, 2R,4S)-4-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 4R)diastereomer

A mixture of 14-O-{[(1R, 2R,5S)-5-tert-butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer and 14-O-{[(1R, 2R,4S)4-tert-butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer (1.12 g, 1.84 mmol) from Example 1 Step A wastreated according to the method of Example 1 Step B. After work up andchromatography of the reaction mixture (silica, ethylacetate/methanol/35% ammonia solution=50/50/1) 14-O-{[(1R, 2R,5S)-5-amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 5R)diastereomer (a) (R_(f)=0.33, 524 mg, 56% yield) and 14-O-{[(1R, 2R,4S)-4-amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 4R)diastereomer (b) (R_(f)=0.22, 160 mg, 17%) were obtained as colorlessamorphous foams.

(a): ¹H NMR (400 MHz, DMSO-d₆, 67, ppm, inter alia): 6.13 (dd, 1H, 19-H,J=11 Hz and 18 Hz), 5.53 (d. 1H, 14-H, J=8 Hz), 5.05 (m, 2H, 20-H), 4.51(d, 1H, 11-OH, J=6 Hz), 3.48 (m, 1H, 2′-H), 3.42 (m, 1H, 11-H),AB-system (ν_(A)=3.37, ν_(B)=3.23, 22-H, J=19 Hz), 2.98 (m, 1H, 1′-H),2.82 (m, 1H, 5′-H), 2.40 (bs, 1H, 4-H), 1.36 (s, 3H, 15-CH₃), 1.06 (s,3H, 18-CH₃), 0.82 (d, 3H, 17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=7 Hz).

(b): ¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.14 (dd, 1H, 19-H,J=11 Hz and 18 Hz), 5.54 (d, 1H, 14-H, J=8 Hz), 5.06 (m, 2H, 20-H), 4.51(bs, 1H, 11-OH), 3.79 (m, 1H, 2′-H), 3.42 (m, 1H, 11-H), AB-system(ν_(A)=3.33, ν_(B)=3.23, 22-H, J=15 Hz), 3.04 (m, 1H, 4′-H), 2.82 (m,1H, 1′-H), 2.40 (bs, 1H, 4-H), 1.37 (s, 3H, 15-CH₃), 1.06 (s, 3H,18-CH₃), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=7 Hz).

Step B. 14-O-{[(1R, 2R,5S)-5-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 5R) diastereomer hydrochloride

14-O-{[(1R, 2R,5S)-5-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 5R)diastereomer (516 mg, 1.02 mmol) was treated according to the method ofExample 1 Step C to obtain 14-O-{[(1R, 2R,5S)-5-amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 5R) diastereomer hydrochloride (533 mg, 96%yield) as colorless amorphous solid.

¹H NMR (500 MHz, DMSO-d₆, δ, ppm, inter alia): 7.7 (bs, 3H, NH₃ ⁺),6.13, 6.12 (2dd, 1H, 19-H, J=11 Hz and 18 Hz), 5.55 (d, 1H, 14-H, J=8Hz), 5.05 (m, 2H, 20-H), 4.53 (d, 1H, 11-OH, J=6 Hz), 3.70 (m, 1H,2′-H), 3.42 (t, 11H, J=6 Hz), 3.35 (m, 2H, 22-H), 3.09 (m, 2H, 1′-H,5′-H), 2.40 (bs, 11H, 4-H), 1.35 (s, 3H, 15-CH₃), 1.05 (s, 3H, 18-CH₃),0.82 (d, 3H, 17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=7 Hz). MS-ESI(m/z): 508 (MH⁺), 530 (MNa⁺, 1015 (2MH⁺), 1037 (2MNa⁺), 542 (MCl⁻).

Example 3 14-O-{[(1R, 2R,4S)-4-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 4R) diastereomer hydrochloride

14-O-{[(1R, 2R,4S)-4-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 4R)diastereomer (152 mg, 0.30 mmol) from Example 2 Step A was treatedaccording to the method of Example 1 Step C to obtain 14-O-{[(1R, 2R,4S)-4-amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 4R) diastereomer hydrochloride (148 mg, 91%yield) as colorless amorphous solid.

¹H NMR (500 MHz, DMSO-d₆, δ, ppm, inter alia): 7.8 (bs, 3H, NH₃ ⁺),6.14, 6.13 (2dd, 1H, 19-H, J=11 Hz and 18 Hz), 5.55 (d, 1H, 14-H, J=8Hz), 5.20 (d, 1H, 2′-OH), 5.05 (m, 2H, 20-H), 4.53 (d, 1H, 11-OH, J=6Hz), 3.88 (m, 1H, 2′-H), 3.42 (t, 1H, 11-H, J=6 Hz), 3.32 (m, 2H, 22-H),3.22 (m, 1H, 4′-H), 2.92 (m, 1H, 1′-H), 2.40 (bs, 1H, 4-4H), 1.35 (s,3H, 15-CH₃), 1.05 (s, 3H, 18-CH₃), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.63(d, 3H, 16-CH₃, J=7 Hz). MS-ESI (m/z): 508 (MH⁺), 530 (MNa⁺), 1015(2MH⁺), 1037 (2MNa⁺), 542 (MCl⁻).

Example 4 14-O-{[(1R, 2R,5R)-5-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 5S)diastereomer

Step A. tert-Butyl-dimethyl-(cis-3,4-epoxycyclohexyloxy)-silane

To a solution of 3-cyclohexen-1-ol (Amburgey, J. C.; Shuey, S. W.;Pedersen, L. G.; Hiskey R., Bioorganic Chemistry 1994, 22, 172-197.) (10g, 102 mmol) in 200 ml of dichloromethane was added vanadylacetylacetonate (0.5 g, cat.) and tert-butyl hydroperoxide (20.4 ml 5.5Min decane, 112 mmol) and stirred overnight at room temperature. Theresulting reaction mixture was treated with tert-butyldimethylsilylchloride (16.9 g, 112 mmol), imidazole (9.02 g, 132 mmol) and4-dimethylaminopyridine (2.49 g, 20 mmol) at 4° C. and stirred for 5hours at room temperature. The reaction mixture was diluted withdichloromethane and subsequently extracted with 10% NaHSO₃ solution,saturated NaHCO₃ solution and brine. The organic layer was dried oversodium sulfate and filtered. The, filtrate was concentrated underreduced pressure and subjected to chromatography (silica,cyclohexane/ethyl acetate=15/1) to yield pure protectedtert-butyl-dimethyl-(cis-3,4-epoxycyclohexyloxy)-silane (R_(f)=0.35,18.3 g, 79% yield) as colorless oil.

¹H NMR (500 MHz, DMSO-d₆, δ, ppm): 3.55 (m, 1H), 3.00 (m, 2H), 2.15 (m,1H), 2.00 (m, 1H), 1.80 (m, 1H), 1.50 (m, 1H), 1.35 (m, 1H), 1.35 (m,1H), 1.25 (m, 1H), 0.83 (s, 9H, tert-butyl), 0.0 (s, 9H, Si(CH₃)₂).

Step B. 14-O-{[(1R, 2R,5S)-5-(tert-Butyl-dimethyl-silyloxy)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer

tert-Butyl-dimethyl-(cis-3,4-epoxycyclohexyloxy)-silane (6.41 g, 28mmol) was treated with pleuromutilin thiol according to the method ofExample 1 Step A2. Crude 14-O-{[(1R, 2R,5S)-5-(tert-butyl-dimethyl-silyloxy)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer was obtained as colorless amorphous foam which wasdirectly used for the next step.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.13 (dd, 1H, 19-H, J=11Hz and 18 Hz), 5.52 (d, 1H, 14-H, J=8 Hz), 5.05 (m, 2H, 20-H), 4.78 (dd,1H, 2′-OH, J=5 Hz and 6 Hz), 4.48 (d, 1H, 11-OH, J=6 Hz), 3.88 (m, 1H,5′-H), 3.15-3.45 (m, 4H, 2′-H, 11-H, 22-CH₂), 2.92 (m, 1H, 1′-H), 2.38(bs, 1H, 4-H), 1.36 (s, 3H, 15-CH₃), 1.05 (s, 3H, 18-CH₃), 0.86 (s, 9H,tert-butyl), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=7 Hz),0.0 (s, 6H, Si(CH₃)₂).

Step C. 14-O-{[(1R, 2R,5S)-2,5-Dihydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 5R)diastereomer

To a solution of 14-O-{[(1R, 2R,5S)-5-(tert-butyl-dimethyl-silyloxy)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer (9.46 g, 15.1 mmol) in 25 ml of THF a mixture ofacetic acid and water (3:1, 100 ml) was added and stirred for 2 days at40° C. The reaction mixture was concentrated nearly to dryness underreduced pressure and the residue was dissolved in ethyl acetate andsubmitted to chromatography (silica, cyclohexane/ethyl acetate=⅓) toyield the 14-O-{[(1R, 2R,5S)-2,5-dihydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 5R)diastereomer (R_(f)=0.27, 7.07 g, 92% yield) as colorless amorphousfoam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.12 (dd, 1H, 19-H, J 11Hz and 18 Hz), 5.53 (d, 1H, 14-H, J=8 Hz), 5.05 (m, 2H, 20-H), 4.72 (dd,1H, 2′-OH, J=2 Hz and 5 Hz), 4.48 (d, 1H, 11-OH, J=6 Hz), 4.43 (t, 1H,5′-OH), 3.68 (m, 1H, 5′-H), 3.45-3.20 (m, 4H, 2′-H, 11-H, 22-H), 2.94(m, 1H, 1′-H), 2.38 (bs, 1H, 4-H), 1.36 (s, 3H, 15-CH₃), 1.06 (s, 3H,18-CH₃), 0.82 (d, 3H, 17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=7 Hz).MS-ESI (m/z): 531 (MNa⁺), 1039 (2MNa⁺).

Step D. 14-O-{[(1R, 2R,5S)-2-Hydroxy-5-methanesulfonyloxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer

To a solution of 14-O-{[(1R, 2R,5S)-2,5-dihydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 5R)diastereomer (6.07 g, 11.9 mmol) in 36 ml of pyridine was addedmethanesulfonyl chloride (1.1 ml, 14.3 mmol) and the resulting mixturewas stirred overnight at room temperature. Subsequently the solvent wasevaporated under reduced pressure; the residue was diluted with 1N HCland extracted three times with ethyl acetate. The combined organiclayers were washed with brine, dried over sodium sulfate and filtered.The filtrate was concentrated and purified by column chromatography(silica, cyclohexane/ethyl acetate=1/1) to yield 14-O-{[(1R, 2R,5S)-2-hydroxy-5-methanesulfonyloxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer (R_(f)=0.15, 2.55 g, 36% yield) as colorlessamorphous foam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.12 (dd, 1H, 19-H, J=11Hz and 18 Hz), 5.54 (d, 1H, 14-H, J=8 Hz), 5.07 (m, 2H, 20-H), 5.00 (t,1H,2′-OH, J=5 Hz), 4.78 (m, 1H, 5′-H), 4.50 (d, 1H, 11-OH, J=6 Hz),3.55-3.25 (m, 4H, 2′-H, 11-H, 22-H)2.91 (m, 1H, 1′-H), 2.38 (bs, 1H,4-H), 1.36 (s, 3H, 15-CH₃), 1.05 (s, 3H, 18-CH₃), 0.80 (d, 3H, 17-CH₃,J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=7 Hz).

Step E. 14-O-{[(1R, 2R,5R)-5-Azido-2-hydroxy-5-methanesulfonyloxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5S) diastereomer

A solution of 14-O-{[(1R, 2R,5S)-2-hydroxy-5-methanesulfonyloxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer (2.55 g, 4.35 mmol) and sodium azide (0.85 g, 13mmol) in 30 ml of dimethylformamide was heated for 6 hours at 80° C. Thereaction mixture was diluted with water and brine and extracted threetimes with ethyl acetate. The combined organic layers were washed withwater and brine, dried over sodium sulfate and filtered. The solvent wasremoved under reduced pressure and crude 14-O-{[(1R, 2R,5R)-5-azido-2-hydroxy-5-methanesulfonyloxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5S) diastereomer (quantitative yield, cyclohexane/ethyl acetate=1/1,R_(f)=0.35) was obtained as amorphous foam which was directly used forthe next step.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.15, 6.13 (2dd, 1H,19-H, J=11 Hz and 18 Hz), 5.56, 5.54 (2d, 1H, 14-H, J=8 Hz), 5.05 (m,2H, 20-H), 4.90 (d, 1H, 2′-OH, J=5 Hz), 4.50, 4,49 (2d, 1H, 11-OH, J=6Hz), 3.50-3.25 (m, 5H, 2′-H, 5′-H, 11-H, 22-H), 2.64 (m, 1H, 1′-H), 2.40(bs, 1H, 4-H), 1.36 (s, 3H, 15-CH₃), 1.06 (s, 3H, 18-CH₃), 0.81 (d, 3H,17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=7 Hz).

Step F. 14-O-{[(1R, 2R,5R)-5-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 5S)diastereomer

Triphenylphosphine (1.18 g, 4.50 mmol) was added to a solution of14-O-{[(1R,2R,5R)-5-azido-2-hydroxy-5-methanesulfonyloxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5S) diastereomer (2.4 g, 4.50 mmol) in 30 ml of THF and stirredovernight at room temperature. Subsequently water (approx. 3 ml) wasadded and the reaction mixture was heated for 1 hour at reflux. Afterevaporation of the solvent the residue was diluted with water and brineand extracted three times with ethyl acetate. The combined organiclayers were dried over sodium sulfate, filtered and subjected tochromatography (silica, ethyl acetate/methanol/35% ammoniasolution=100/100/1),) to yield 14-O-{[(1R, 2R,5R)-5-amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 5S)diastereomer (R_(f)=0.3, 1.74 g, 79% yield) as colorless amorphous foam.

¹H NMR (400 MHz, DMSO-d6, δ, ppm, inter alia): 7.25, 6.65 (2bs, 1H, NH),6.14 (dd, 1H, 19-H, J=11 Hz and 18 Hz), 5.54 (d, 1H, 14-H, J=8 Hz), 5.04(m, 2H, 20-H), 4.50 (bs, 1H, 11-OH), 3.55-3.10 (m, 5H, 2′-H, 5′-H, 11-H,22-H), 2.58 (m, 1H, 1′-H), 2.40 (bs, 1H, 4-H), 1.36 (s, 3H, 15-CH₃),1.05 (s, 3H, 18-CH₃), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.62 (d, 3H, 16-CH₃,J=7 Hz). MS-ESI (m/z): 508 (MH⁺), 530 (MNa⁺), 1037 (2MNa⁺).

Example 5 14-O-{[(1R, 2R,3R)-3-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 3S) diastereomer hydrochloride

Step A. 14-O-{[(1R, 2R,3R)-3-tert-Butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3S) diastereomer

(cis)-2,3-Epoxycyclohexyl-carbamic acid tert-butyl ester (O'Brien, P.;Childs, A., C.; Ensor, G. Organic Letters 2003, 5(26), 4955-4957.) (1 g,4.69 mmol) was treated with pleuromutilin thiol according to the methodof Example 1 Step A1. After work up and chromatography of the reactionmixture (silica, cyclohexane/ethyl acetate=1/1) 14-O-{[(1R, 2R,3R)-3-tert-butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3S) diastereomer (R_(f)=0.5, 1.32 g, 46%) was obtained as colorlessamorphous foam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.12 (m, 2H, NH, 19 -Hz),5.54 (d, 1H, 14-H, J=8 Hz), 5.05 (m, 2H, 20-H), 4.96 (d, 1H, 2′-OH, J=4Hz), 4.50, 4.99 (2d, lH, 1-OH, J=6 Hz)), 3.65 (m, 1H, 2′-H), 3.57 (m,1H, 3′-H), 3.42 (t, 1H, 11-H, J=6 Hz), AB-system (ν_(A)=3.30, 3.29,ν_(B)=3.23, 3.22, 22-H, J=15 Hz), 3.06 (m, 1H, 1′-1), 2.40 (bs, 1H,4-H), 1.36 (s, 12H, tert-butyl, 15-CH₃), 1.05 (s, 3H, 18-CH₃), 0.82 (d,3H, 17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=6 Hz).

Step B. 14-O-{[(1R, 2R,3R)-3-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 3S)diastereomer

14-O-{[(1R, 2R,3R)-3-tert-Butoxycarbonylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3S) diastereomer (400 mg, 0.658 mmol) was treated according to themethod of Example 1 Step B. After work up and chromatography of thereaction mixture (silica, ethyl acetate/methanol=1/5) 14-O-{[(1R, 2R,3R)-3-amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 3S)diastereomer (R_(f)=0.1, 249 mg, 75%) was obtained as colorlessamorphous foam.

-   MS-ESI (m/z): 508 (MH⁺), 530 (MNa⁺), 1015 (2MH⁺), 1037 (2MNa⁺).    Step C. 14-O-{[(1R, 2R,    3R)-3-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin    hydrochloride+(1S, 2S, 4S) diastereomer hydrochloride

14-O-{[(1R, 2R,3R)-3-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 3S)diastereomer (249 mg, 0.49 mmol) was treated according to the method ofExample 1 Step C to obtain 14-O-{[(1R, 2R,3R)-3-amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 3S) diastereomer hydrochloride (247 mg, 93%yield) as colorless amorphous solid.

¹H NMR (400 Mhz, DMSO-d₆, δ, ppm, inter alia): 7.8 (bs, 3H, NH₃ ⁺), 6.13(d, 2H, 19-Hz, J=11 Hz and 18 Hz), 5.80 (d, 1H, 2′-OH, J=4 Hz), 5.55 (d,1H, 14-H, J=8 Hz), 5.05 (m, 2H, 20-H), 4.55, 4.54 (2d, 1H, 11-OH, J=6Hz)), 3.87 (m, 1H, 2′-H), 3.42 (t, 1H, 11-H, J=6 Hz), AB-system(ν_(A)=3.35, ν_(B)=3.24, 22-H, J=15 Hz), 3.20, 3.13 (2m, 1H, 3′-H,1′-H), 2.40 (bs, 1H, 4-H), 1.36 (s, 3H, 15-CH₃), 1.05 (s, 3H, 18-CH₃),0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=7 Hz). MS-ESI(m/z): 508 (MH⁺), 1015 (2MH⁺), 542 (MCl⁻).

Example 6 14-O-{[(1R, 2R,4R)-4-Diethylamino-2-hydroxy-cyclobexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4S) diastereomer and 14-O-{[(1R, 2R,4R)-4-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S,4S) diastereomer

To a solution of 14-O-{[(1R, 2R,4R)-4-amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 4S)diastereomer (900 mg, 1.77 mmol) from Example 1 Step B in 10 mldichloromethane was added acetaldehyde (2.77 ml, 1M in dichloromethane)and acetic acid (77 μl, 1.77 mmol) and stirred for 30 minutes at roomtemperature. The resulting reaction mixture was treated with sodiumtriacetoxyborohydride (750 mg, 3.54 mmol) and stirred overnight at roomtemperature, diluted with dichloromethane and subsequently extractedwith NaHCO₃ solution and brine. The organic layer was dried over sodiumsulfate and filtered. The filtrate was subjected to chromatography(silica, ethyl acetate/methanol/35% ammonia solution=50/50/1) to yield14-O-{[(1R, 2R,4R)-4-diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4S) diastereomer (a) (92 mg, 9% yield) and 14-O-{[(1R, 2R,4R)-4-ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S,4S) diastereomer (b) (163 mg, 17% yield) as colorless amorphous foams.

(a): ¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.14 (dd, 1H, 19-H,J=11 Hz and 18 Hz), 5.55 (d, 1H, 14-H, J=8 Hz), 5.05 (m, 2H, 20-H), 4.83(d, 1H, 2′-OH, J=4 Hz), 4.47 (d, 1H, 11-OH, J=6 Hz), 3.42 (m, 1H, 11-H),AB-system (ν_(A)=3.50, 3.42, ν_(B)=3.30 ,3.27, 22-H, J=15 Hz), 3.25 (m,1H, 2′-H), 2.50 (m, 2H, 1′-H, 4′-H), 2.40 (m, 5H, NCH₂, 4-H), 1.36 (s,3H, 15-CH₃), 1.05 (s, 3H, 18-CH₃), 0.92 (t, 6H, NCH₂CH₃, J=7 Hz), 0.81(d, 3H, 17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=7 Hz). MS-ESI (m/z): 564(MH⁺), 586 (MNa⁺), 562 (M-H)⁻.

(b): ¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.13 (dd, 1H, 19-H,J=11 Hz and 18 Hz), 5.54 (d, 1H, 14-H, J=8 Hz), 5.05 (m, 2H, 20-H), 4.48(d, 1H, 11-OH, J=6 Hz), 3.42 (m, 1H, 11-H), AB-system (ν_(A=)3.48,ν_(B)=3.25,22-H, J=15 Hz), 2.55 (m, 2H, 1′-H, 4′-H), 2.40 (bs, 1H, 4-H),1.35 (s, 3H, 15-CH₃), 1.05 (s, 3H, 18-CH₃), 0.95 (t, 3H, NCH₂CH₃, J=7Hz), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=6 Hz). MS-ESI(m/z): 536 (MH⁺), 558 (MNa⁺), 534 (M-H)⁻.

Example 7 14-O-{[(1R, 2R,5S)-5-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 5R) diastereomer hydrochloride

Step A. N-Ethyl-N-(cyclohex-3-enyl)-carbamic acid tert-butyl ester

To a solution of cyclohex-3-enyl-carbamic acid tert-butyl ester(Kampferer, P.; Vasella, A. Helvetica Chimica Acta 2004, 87, 2764-2789)(4.34 g, 22 mmol) in 20 ml of DMSO was added sodium hydride (880 mg, 60%dispersion, 22 mmol) and after one hour of stirring ethyl iodide (1.78ml, 22 mmol). After further stirring for 2 hours at room temperature thereaction mixture was concentrated under reduced pressure. The residuewas diluted with water and brine and extracted three times with ethylacetate. The organic layers were dried over sodium sulfate and filtered.The filtrate was concentrated under reduced pressure and afterchromatography (silica, cyclohexane/ethyl acetate=12/1) the titlecompound (R_(f)=0.30, 2.88 g, 58% yield) was obtained as colorlesssolid.

¹H NMR (400 MHz, CDCl₆, δ, ppm): 5.61 (m, 2H, double bond), 4.08 (bs,1H, NCH), 3.15 (m, 2H, NCH₂), 2.15, 1.75 (2m, 6H), 1.47 (s, 9H,tert-butyl), 1.13 (t, 3H, NCH₂CH₃, J=7 Hz).

Step B. N-ethyl-N-(cis-3,4-epoxycyclohexyl)-carbamic acid tert-butylester

N-Ethyl-N-(cyclohex-3-enyl)-carbamic acid tert-butyl ester (2.87 g, 12.7mmol) was dissolved in 75 ml of dichloromethane and treated with3-chloroperbenzoic acid (4.50 g, 70%, 19 mmol). After stirring at roomtemperature for 20 hours the reaction mixture was concentrated underreduced pressure. The residue was diluted with ethyl acetate andsubsequently extracted with 10% NaHSO₃ solution, saturated NaHICO₃solution and brine. The organic layer was dried over sodium sulfate andfiltered. The filtrate was concentrated under reduced pressure and afterchromatography (silica, cyclohexane/dioxane 5/1) the title compound(R_(f)=0.2, 1.50 g, 49% yield) was obtained.

¹H NMR (400 MHz, CDCl₆, δ, ppm): 4.0 (bs, 1H, NCH), 3.14 (m, 2H, NCH₂),3.06 (bs, 2H, epoxide), 2.13, 2.08, 1.88, 1.60, 1.36 (4m, 6H), 1.47 (s,9H, tert-butyl), 0.08 (t, 3H, NCH₂CH₃).

Step C. 14-O-{[(1R, 2R,5S)-5-(tert-Butoxycarbonyl-ethyl-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer

N-Ethyl-N-(cis-3,4-epoxycyclohexyl)-carbamic acid tert-butyl ester (1.5g, 6.2 mmol) was treated with pleuromutilin thiol according to themethod of Example 1 Step A1. After work up and chromatography of thereaction mixture (silica, cyclohexane/dioxane=3/1) 14-O-{[(1R, 2R,5S)-5-(tert-Butoxycarbonyl-ethyl-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer (R_(f)=0.4, 2.57 g, 65% yield) was obtained ascolorless amorphous foam.

MS-ESI (m/z): 536 (MH⁺), 558 (MNa⁺), 534 (M-H)⁻.Step D. 14-O-{[(1R, 2R,5S)-5-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S,5R) diastereomer

14-O-{[(1R, 2R,5S)-5-(tert-Butoxycarbonyl-ethyl-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer (2.57 g, 4.04 mmol) was treated according to themethod of Example 1 Step B. After work up and chromatography of thereaction mixture (silica, ethyl acetate/methanol/35% ammoniasolution=100/100/1) 14-O-{[(1R, 2R,5S)-5-ethylamino-2-hydroxy-cyclohexylsulfanyl]acetyl}-mutilin+(1 S, 2S,5R) diastereomer (R_(f)=0.3, 1.08 g, 50%) was obtained as colorlessamorphous foam.

¹H NMR (500 MHz, CDCl₆, δ, ppm, inter alia): 6.48 (dd, 1H, 19-H, J=10 Hzand 18 Hz), 5.77 (m, 1H, 14-H), 5.36 (m, 1H, 20-H), 5.22 (d, 1H, 20-H,J=17 Hz), 3.45 (d, 1H, 2′-H), 3.37 (d, 1H, 11-H, J=6 Hz), 3.25 (m, 1H,22-H), 2.97 (m, 1H, 1′-H), 2.91 .(m, 1H, 5′-H), 2.63 (q, 2H, NCH₂, J=7Hz), 2.10 (bs, 1H, 4-H), 1.46 (s, 3H, 15-CH₃), 1.18 (s, 3H, 18-CH₃),1.12 (t, 3H, NCH₂CH₃, J=7 Hz), 0.98 (d, 3H, 17-CH₃, J=7 Hz), 0.73 (d,3H, 16-CH₃, J=7 Hz).

Step E. 14-O-{[(1R, 2R,5S)-5-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 5R) diastereomer hydrochloride

14-O-{[(1R, 2R,5S)-5-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S,5R) diastereomer (86 mg, 0.16 mmol) was treated according to the methodof Example 1 Step C to obtain 14-O-{[(1R, 2R,5S)-5-ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 5R) diastereomer hydrochloride (83 mg, 90% yield)as colorless amorphous solid.

¹H NMR (400 MHz, CDCl₆, δ, ppm, inter alia): 9.3 (bs, 2H, NH₂ ⁺), 6.45(m, 1H, 19-H), 5.73 (d, 1H, 14-H, J=10 Hz), 5.35 (m, 1H, 20-H), 5.22 (d,1H, 22-H, J=18H), 3.85 (m, 1H, 2′-H), 3.33 (m, 3H, 11-H, 22-H), 3.07 (m,2H, NCH₂), 2.10 (bs, 1H, 4-H), 1.50 (t, 3H, NCH₂CH₃, J=7 Hz), 1.45 (s,3H, 15-CH₃), 1.18 (s, 3H, 18-CH₃), 0.90 (d, 3H, 17-CH₃, J=7 Hz), 0.73(d, 3H, 16-CH₃, J=7 Hz). MS-ESI (m/z): 536 (MH⁺), 570 (MCl⁻).

Example 8 14-O-{[(1R, 2R,5S)-5-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 5R) diastereomer hydrochloride

Step A. 14-O-{[(1R, 2R,5S)-5-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilia+(1S,2S, 5R) diastereomer

14-O-{[(1R, 2R,5S)-5-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S,5R) diastereomer (250 mg, 0.47 mmol) from Example 7 Step D was treatedwith acetaldehyde (53 μl, 0.93 mmol) according to the method of Example6. After work up and chromatography of the reaction mixture (silica,ethyl acetate/methanol=1/1) 14-O-{[(1R, 2R,5S)-5-diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer (R_(f)=0.2, 230 mg, 87%) was obtained as colorlessamorphous foam.

¹H NMR (400 MHz, CDCl₆, δ, ppm, inter alia): 6.48 (dd, 1H, 19-H, J=11 Hzand 17 Hz), 5.77 (d, 1H, 14-H, J=8 Hz), 5.36 (m, 1H, 20-H), 5.22 (d, 1H,20-H, J=17 Hz), 3.57, 3.36, 3.21, 3.03, 2.72 (5m, 6H, 1′-H, 2′-H, 5′-H,11-H, 22-H), 2.59 (m, 4H, NCH₂), 2.11 (bs, 1H, 4-H), 1.46 (s, 3H,15-CH₃), 1.18 (s, 3H, 18-CH₃), 0.98 (t, 6H, NCH₂CH₃, J=7 Hz), 0.88 (d,3H, 17-CH₃, J=7 Hz), 0.73 (d, 3H, 16-CH₃, J=7 Hz).

Step B. 14O-{[(1R, 2R,5S)-5-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 5R) diastereomer hydrochloride

14-O-{[(1R, 2R,5S)-5-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer (230 mg, 0.41 mmol) was treated according to themethod of Example 1 Step C to obtain 14-O-{[(1R, 2R,5S)-5-diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinhydrochloride+(1S, 2S, 5R) diastereorner hydrochloride (223 mg, 91%yield) as colorless amorphous solid.

¹H NMR (400 MHz, CDCl₆, δ, ppm, inter alia): 11.5 (bs, 3H, NH⁺), 6.46(dd, 1H, 19-H, J=11 Hz and 17 Hz), 5.74 (d, 1H, 14-H, J=8 Hz), 5.34 (m,1H, 20-H), 5.22 (d, 1H, 22-H, J=17H), 3.98 (m, 1H, 2′-H), 3.60-2.90 (m,9H, 1′-H, 5′-H, 11-H, 22-H, NCH₂), 2.10 (bs, 1H, 4-H), 1.48 (m, 9H,NCH₂CH₃, 15-CH₃), 1.18 (s, 3H, 18-CH₃), 0.89 (d, 3H, 17-CH₃, J=7 Hz),0.73 (d, 3H, 16-CH₃, J=7 Hz). MS-ESI (m/z): 564 (MH⁺), 586 (MNa⁺), 1149(2MNa⁺), 598 (MCl⁻).

Example 9 14-O-{[(1R, 2R,4S)-4-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer

14-O-{[(1R, 2R, 4S)-4-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 4R) diastereomer (680 mg, 1.34 mmol) from Example 2Step A was treated according to the method of Example 6. After work upand chromatography of the reaction mixture (silica, ethylacetate/methanol=1/1) 14-O-{[(1R, 2R,4S)-4-diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer (R_(f)=0.2, 129 mg, 17%) was obtained as colorlessamorphous foam.

¹H NMR (500 MHz, DMSO-d₆, δ, ppm, inter alia): 6.13 (dd, 1H, 19-H, J=11Hz and 18 Hz), 5.54 (d, 1H, 14-H, J=8 Hz), 5.05 (m, 2H, 20-H), 4.70 (d,1H, 2′-OH), 4.49 (d, 1H, 11-OH, J=6 Hz), 3.70 (m, 1H, 2′-H), 3.42 (t,1H, 11-H, J=6 Hz), AB-system (ν_(A)=3.36, ν_(B)=3.22, 22-H, J=15 Hz),2.72 (m, 2H, 1′-H, 5′-H), 2.47 (m, 2H, NCH₂), 2.40 (bs, 1H, 4-H), 1.35(s, 3H, 15-CH₃), 1.04 (s, 3H, 18-CH₃), 0.95 (t, 3H, NCH₂CH₃, J=7 Hz),0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.62 (d, 3H, 16-CH₃, J=7 Hz). MS-ESI(m/z): 536 (MH⁺), 558 (MNa⁺), 534 (M-H)⁻.

Example 10 14-O-{[(1R, 2R,5R)-5-Diethylamino-2-hydroxy-cyclohexylsuffanyl]-acetyl}-mutilin+(1S,2S, 5S) diastereomer

14-O-{[(1R, 2R,5R)-5-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 5S)diastereomer (420 mg, 0.827 mmol) from Example 4 Step F was treatedaccording to the method of Example 6. After work up and chromatographyof the reaction mixture (silica, ethyl acetate/methanol/35% ammoniasolution=50/50/1) 14-O-{[(1R, 2R,5R)-5-diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5S) diastereomer (R_(f)=0.2, 95 mg, 20%) was obtained as colorlessamorphous foam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.13 (dd, 1H, 19-H, J=11Hz and 18 Hz), 5.55 (d, 1H, 14-H, J=8 Hz), 5.05 (m, 2H, 20-H), 4.79 (m,1H, 2′-OH), 3.55-3.15 (m, 2′-H, 5′-H, 11-H, 22-H), 2.58 (m, 1H, 1′-H),2.40 (m, 5H, NCH₂, 4-H), 1.37 (s, 3H, 15-CH₃), 1.06 (s, 3H, 18-CH₃),0.92 (t, 3H, NCH₂CH₃, J=7 Hz), 0.83 (d, 3H, 17-CH₃, J=7 Hz), 0.63 (d,3H, 16-CH₃, J=7 Hz). MS-ESI (m/z): 564 (MH⁺), 586 (MNa⁺), 562 (M-H)⁻,598 (MCl⁻).

Example 11 14-O-{[(1R, 2R,3R)-3-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S,3S) diastereomer

Step A. [(1R, 2R,3R)-2-Hydroxy-3-(2,4,6-trimethyl-benzylsulfanyl)-cyclobexyl]-carbamicacid tert-butyl ester+(1S, 2S, 3S) diastereomer

To a solution of (cis)-2,3-epoxycyclohexyl-carbamic acid tert-butylether (O'Brien, P.; Childs, A., C.; Ensor, G. Organic Letters 2003,5(26), 4955- 4957.) (14.9 g, 68.9 mmol) and 2,4,6-trimethylbenzylmercaptan (11.5 g, 68.9 mmol) in 50 ml of methanol was added 10N NaOH (5ml, 50 mmol) and the resulting mixture was stirred for 16 hours at roomtemperature. The reaction mixture was diluted with water and brine andextracted with ethyl acetate three times. The organic layers were driedover sodium sulfate and filtered. The filtrate was subjected tochromatography (silica, cyclohexane/ethyl acetate=5/1) to yield [(1R,2R,3R)-2-hydroxy-3-(2,4,6-trimethyl-benzylsulfanyl)-cyclohexyl]-carbamicacid tert-butyl ester+(1S, 2S, 3S) diastereomer (R_(f)=0.25, 5.92g, 23%yield) as colorless amorphous foam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm): 6.78 (s, 2H, aromat.-H), 6.15 (bd,OH), 4.95 (bd, NH), 3.75 (d, 1H, SCH₂), 3.68 (m, 2H), 3.02 (m, 1H, SCH),2.30 (s, 9H, CH₃), 2.30, 1.90, 1.40 (3m, 6H), 1.35 (s, 9H, tert-butyl).

Step B. [(1R, 2R,3R)-2-(tert-Butyl-dimethyl-silanyloxy)-3-(2,4,6-trimethyl-benzylsulfanyl)-cyclohexyl]-carbamicacid tert-butyl ester+(1S, 2S, 38) diastereomer

A solution of [(1R, 2R,3R)-2-hydroxy-3-(2,4,6-trimethyl-benzylsulfanyl)-cyclohexyl]-carbamicacid tert-butyl ester+(1S, 2S, 3S) diastereomer (2.46 g, 6.49 mmol) in50 ml of dimethylformamide was treated with tert-butyldimethylsilylchloride (978 mg, 6.49 mmol) and imidazole (552 mg, 8.11 mmol) andstirred at 80 ° C. for 5 days. The reaction mixture was concentratedunder reduced pressure. The residue was diluted with 0.1 N HCl andextracted three times with ethyl acetate. The combined organic layerswere washed with brine, dried over sodium sulfate and filtered. Afterchromatography (silica, cyclohexane/ethyl acetate=10/1) [(1R, 2R,3R)-2-(tert-butyl-dimethyl-silanyloxy)-3-(2,4,6-trimethyl-benzylsulfanyl-cyclo-hexyl]-carbamicacid tert-butyl ester+(1S, 2S, 3S) diastereomer (R_(f)=0.25, 3.0 g, 94%yield) was obtained.

¹H NMR (500 MHz, DMSO-d6, δ, ppm): 6.80 (s, 2H, aromat-H), 6.20 (bd,NH), 3.90, 3.75, 3.63 (3m, 4H, NCH, OCH, SCH₂), 2.98 (m, 1H, SCH), 2.30(s, 9H, CH₃), 1.90, 1.50, 1.33 (3m, 6H), 1.35. (s, 9H, C-tert-butyl),0.85 (s, 9H, Si-tert-butyl), 0.0 (s, 6H, Si(CH₃)₂).

Step C. [(1R, 2R,3R)2-(tert-Butyl-dimethyl-silanyloxy)-3-(2,4,6trimethyl-benzylsulfanyl)-cyclohexyl]-ethyl-carbamicacid tert-butyl ester+(1S, 2S, 3S) diastereomer

To a solution of [(1R, 2R,3R)-2-(tert-butyl-dimethyl-silanyloxy)-3-(2,4,6-trimethyl-benzylsulfanyl)-cyclohexyl]-carbamicacid tert-butyl ester+(1S, 2S, 3S) diastereomer (3.0 g, 6.08 mmol) wastreated with ethyl iodide according to the method of Example 7 Step A.After work up and chromatography of the reaction mixture (silica,cyclohexane / ethyl acetate=3/1) [(1R, 2R,3R)-2-(tert-butyl-dimethyl-silanyloxy)-3-(2,4,6-trimethyl-benzylsulfanyl)-cyclohexyl]-ethyl-carbamicacid tert-butyl ester+(1S, 2S, 3S) diastereomer (1.20 g, 38%) wasobtained.

MS-ESI (m/z): 544 (MNa⁺), 1065 (2MNa⁺).Step D. [(1R, 2R,3R)-2-(tert-Butyl-dimethyl-silanyloxy)-3-mercapto-cyclohexyl]-ethyl-carbamicacid tert-butyl ester+(1S, 2S, 3S) diastereomer

A solution of [(1R, 2R,3R)-2-(tert-butyl-dimethyl-silanyloxy)-3-(2,4,6-trimethyl-benzylsulfanyl)-cyclohexyl]-ethyl-carbamicacid tert-butyl ester+(1S, 2S, 3S) diastereomer (1.20 g, 2.30 mmol) in10 ml of tetrahydrofuran and 20 ml of liquid ammonia was treated at −78°C. under an argon atmosphere with sodium (106 mg, 4.60 mmol) and stirredat −78° C. for one hour. Then solid ammonium chloride was added and thereaction mixture was warmed to room temperature, diluted withtetrahydrofuran and flushed with nitrogen. The residual mixture wasfiltered and concentrated under reduced pressure to yield crude [(1R,2R,3R)-2-(tert-butyl-dimethyl-silanyloxy)-3-mercapto-cyclohexyl]-ethyl-carbamicacid tert-butyl ester+(1S, 2S, 3S) diastereomer (quantitative yield)which was directly used for the next step.

MS-ESI (m/z): 412 (MNa⁺), 801 (2MNa⁺).Step E. 14-O-{[(1R, 2R,3R)-3-(tert-Butoxycarbonyl-ethyl-amino)-2-(tert-butyl-dimethyl-silanyloxy)-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3S) diastereomer

Crude [(1R, 2R,3R)-2-(tert-butyl-dimethyl-silanyloxy)-3-mercapto-cyclohexyl]-ethyl-carbamicacid tert-butyl ester+(1S, 2S, 3S) diastereomer (895 mg, 2.30 mmol) wasdissolved in 30 ml of tetrahydrofuran and treated subsequently withpleuromutilin tosylate (979 mg, 1.84 mmol) and potassium tert-butoxide(206 mg, 1.84 mmol) and the resulting mixture was stirred for 16 hoursat room temperature. After evaporation of the solvent the residue wasdiluted with 1N HCl and extracted three times with ethyl acetate. Thecombined organic layers were washed with NaHCO₃ solution and brine,dried over sodium sulfate and filtered. After chromatography (silica,cyclohexane/ethyl acetate=10/1) 14-O-{[(1R, 2R,3R)-3-(tert-butoxycarbonyl-ethyl-amino)-2-(tert-butyl-dimethyl-silanyloxy)-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3S) diastereomer (R_(f)=0.5, 468 mg, 27% yield) was obtained.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.14 (dd, 1H, 19-H, J=1Hz and 18 Hz), 5.55 (d, 1 H, 14-H, J=8 Hz), 5.05 (m, 2H, 20-H), 4.49 (m,1H, 11-OH), 3.94 (m, 1H, 2′-H), 3.43 (t, 1H, 11-OH, J=6 Hz), 3.28, 3.04(2m, 7H, 1′-H, 3′-H, 11-H, 22-H, NCH₂), 2.40 (bs, 1H, 4-H), 1.40 (s, 9H,tert-butyl), 1.36, 1.35 (2s, 3H, 15-CH₃), 1.05 (s, 3H, 18-CH₃), 0.87 (s,9H, Si-tert-butyl), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.64, 0.62 (2d, 3H,16-CH₃, J=7 Hz) 0.05,-0.05 (2s, 6H, Si(CH₃)₂).

Step F. 14-O-{[(1R, 2R,3R)-3-Etlhylamino-2-(tert-butyl-dimethyl-silanyloxy)-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3S) diastereomer and 14-O-([(1R, 2R,3R)-3-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S,3S) diastereomer

14-O-{[(1R, 2R,3R)-3-(tert-Butoxycarbonyl-ethyl-amino)-2-(tert-butyl-dimethyl-silanyloxy)-cyclohexylsulfanyl]-acetyl)-mutilin+(1S,2S, 3S) diastereomer (468 mg, 0.624 mmol) was treated withtrifluoroacetic acid overnight according to the method of Example 1 StepB. After work up and chromatography of the reaction mixture (silica,ethyl acetate/methanol=1/2) 14-O-{[(1R, 2R,3R)-3-ethylamino-2-(tert-butyl-dimethyl-silanyloxy)-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3S) diastereomer (a) (R_(f)=0.6, 144 mg, 36% yield) and 14-O-([(1R,2R, 3R)-3-ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl} -mutilin+(1S,2S, 3S) diastereomer (b) (R_(f)=0.25, 177 mg, 53% yield) were obtainedas colorless solids.

(a): MS-ESI (m/z): 672 (MNa⁺), 1321 (2MNa⁺).

(b): ¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.14 (dd, 1H,19-H,J=11 Hz and 18 Hz), 5.54 (d, 1H, 14-H, J=8 Hz), 5.05 (m, 2H, 20-H),4.76 (m, 1H, 2′-OH), 4.49, 4.48 (2d, 1H, 11-OH, J=6 Hz), 3.55 (m, 1H,2′-H), 3.42 (t, 1H, 11-H, J=6 Hz), AB-system (ν_(A)=3.37, ν_(B)=3.18,22-H, J=15 Hz), 3.05 (m, 1H, 3′-H), 2.66 (m, 1H, 1′-H), 2.50 (m, 2H,NCH₂), 2.40 (bs, 1H, 4-H), 1.36 (s, 3H, 15-CH₃), 1.05 (s, 3H, 18-CH₃),0.98 (t, 3H, NCH₂CH₃, J=7 Hz), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.62 (d,3H, 16-CH₃, J=7 Hz). MS-ESI (m/z): 536 (MH⁺), 558 (MNa⁺), 1071 (2MH⁺),1093 (2MNa⁺), 534 (M-H)⁻.

Example 12 14-O-{[(1R, 2R,3R)-3-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3S) diastereomer

Step A. 14-O-{[(1R, 2R,3R)-3-Diethylamino)-2-(tert-butyl-dimethyl-silanyloxy)-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 35) diastereomer

14-O-{[(1R, 2R,3R)-3-Ethylamino-2-(tert-butyl-dimethyl-silanyloxy)-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3S) diastereomer (144 mg, 0.222 mmol) from Example 11 Step F wastreated with acetaldehyde (25 μl, 0.444 mmol) according to the method ofExample 6. After work up and chromatography of the reaction mixture(silica, ethyl acetate/methanol=2/1) 14-O-{[(1R, 2R,3R)-3-diethylamino)-2-(tert-butyl-dimethyl-silanyloxy)-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3S) diastereomer (R_(f)=0.5, 110 mg, 73%) was obtained as colorlessamorphous foam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.14 (dd, 1H, 19-H, J=11Hz and 18 Hz), 5.53 (d, 1H, 14-H, J=7 Hz), 5.05 (m, 2H, 20-H), 4.98 (m,1H, 11-OH), 3.97 (m, 1H, 2′-H), 3.42 (t, 1H, 11-H, J=6 Hz), 3.24 (m, 2H,H-22), 3.00 (m, 1H, 1′-H), 2.70 (m, 1H, 3′-H), 2.55 (m, 4H, NCH₂), 2.40(bs, 1H, 4-H), 1.36 (s, 3H, 15-CH₃), 1.05 (s, 3H, 18-CH₃), 0.87 (m, 15H,NCH₂CH₃, Si-tert-butyl), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.62, 0.60 (2d,3H, 16-CH₃, J=7 Hz), 0.07 (s, 6H, Si(CH₃)₂).

Step B. 14-O-{[(1R, 2R,3R)-3-Diethylamino-2-hydroxy-cyclohexylsulfanyll-acetyl}-mutilin+(1S,2S, 3S) diastereomer

To a solution of 14-O-{[(1R, 2R,3R)-3-diethylamino-2-(tert-butyl-dimethyl-silanyloxy)-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3S) diastereomer (101 mg, 0.149 mmol) in 5 ml of tetrahydrofuran wasadded tetrabutylammonium fluoride (0.44 ml, 1M in THF, 0.447 mmol).After stirring at room temperature for 2 days the reaction mixture wasconcentrated under reduced pressure, diluted with NaHCO₃ solution andextracted two times with ethyl acetate. The combined organic layers weredried over sodium sulfate and filtered. The filtrate was subjected tochromatography (silica, ethyl acetate/methanol=1/2) to obtain14-O-{[(1R, 2R,3R)-3-diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3S) diastereomer (R_(f)=0.2, 8 mg, 10% yield) as colorless amorphousfoam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.15, 6.14 (2dd, 1H,19-H, J=11 Hz and 18 Hz), 5.54 (d, 1H, 14-H, J=8 Hz), 5.05 (m, 2H,20-H), 4.99, 4.42 (2m, 2H, 11-OH, 2′-OH), 3.87 (m, 1H, 2′-H), 3.42 (t,1H, 11-H, J=6 Hz), 3.25 (m, 2H, H-22), 3.05 (m, 1H, 1,′-H), 2.60 (m, 3H,3′-H, NCH₂), 2.40 (bs, 1n, 4-H), 1.35 (s, 3H, 15-CH₃), 1.05 (s, 3H,18-CH₃), 0.90 (m, 6H, NCH₂CH₃), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.62, 0.61(2d, 3H, 16-CH₃, J=7 Hz). MS-E3SI (m/z): 564 (MH⁺), 586 (MNa⁺), 1149(2MNa⁺), 562 (M-H)⁻.

Example 13 14-O-{[(1R, 2R,4S)-4-(Formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer

Step A. 14-O-{[(7R,8R)-7-Hydroxy-1,4-dioxa-spiro[4.51dec-8-ylsulfanyl]-acetyl}-mutilin+(7S,8S) diastereomer

7,8-Epoxy-1,4-dioxa-spiro[4.5]decane (Zhang, L.; Koreeda, M. OrganicLetters 2002, 4(21), 3755-3758.) (6.25 g, 40 mmol) and pleuromutilinthiol (8 g, 20 mmol) were treated according to the method of Example 1Step A2. After work up and chromatography of the reaction mixture(silica, cyclohexane/ethyl acetate=1/1) 14-O-{[(7R,8R)-7-hydroxy-1,4-dioxa-spiro[4.5]dec-8-ylsulfanyl]-acetyl}-mutilin+(7S,8S) diastereomer (R_(f)=0.3, 8.40 g, 76% yield) was obtained ascolorless amorphous foam.

¹H NMR (500 MHz, DMSO-d₆, δ, ppm, inter alia): 6.14 (dd, 1H, 19-H, J 11Hz and 18 Hz), 5.55 (d, 1H, 14-H, S=8 Hz), 5.05 (m, 2H, 20-H), 4.95 (d,1H, 2′-OH, J 6 Hz), 4.50 (d, 1H, 11-OH, J=6 Hz), 3.82 (m, 4H, OCH₂CH₂O),3.55-3.25 (m, 4H, 2′-H, 11-H, H-22), 2.58 (mn, 1H, 1′-H), 2.40 (bs, 1H,4-H), 1.35, 1.34 (2s, 3H1, 15-CH₃), 1.05 (s, 3H, 18-CH₃), 0.81 (d, 3H,17-CH₃, J=7 Hz), 0.62 (d, 3H, 16-CH₃, J=7 Hz). MS-ESI (m/z): 573 (MNa⁺),549 (M-H)⁻.

Step B. 14-O-{[(7R,8R)-7-(tert-Butyl-diphenyl-silanyloxy)-1,4-dioxa-spiro[4.5]dec-8-ylsulfanyl]-acetyl}-mutilin+(7S,8S) diastereomer

A solution of 14-O-{[(7R,8R)-7-hydroxy-1,4-dioxa-spiro[4.5]dec-8-ylsulfanyl]-acetyl}-mutilin+(7S,8S) diastereomer (8.4 g, 15.3 mmol) in 50 ml of dimethylformamide wastreated with tert-butyldiphenylsilyl chloride (5.16 ml, 19.8 mmol) andimidazole (1.66 g, 24.4 mmol) and stirred overnight at 80 ° C. Thereaction mixture was concentrated under reduced pressure. The residuewas diluted with water and brine and extracted with ethyl acetate threetimes. The combined organic layers were washed with water and brine,dried over sodium sulfate and filtered. After chromatography (silica,cyclohexane/ethyl acetate=1/1) 14-O-{[(7R,8R)-7-(tert-butyl-diphenyl-silanyloxy)-1,4-dioxa-spiro[4.5]dec-8-ylsulfanyl]-acetyl}-mutilin+(7S,8S) diastereomer (R_(f)=0.7, 8.03 g, 67% yield) was obtained.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 7.7-7.35 (m, 10H,aromat,-H), 6.15, 6.13 (2dd, 1H, 19-H, J=11 Hz and 18 Hz), 5,57, 5.53(2d, 1H, 14-H, J=7 Hz), 5.05 (m, 2H, 20-H), 4.50 (m, 1H, 11-OH), 3.30(m, 1H, 2′-H), 3.70-2.80 (m, 8H, OCH₂CH₂O, 1′-H, 11-H, 22-H), 2.40 (bs,1H, 4-H), 1.39, 1.36 (2s, 3H, 15-CH₃), 1.05 (s, 3H, 18-CH₃), 1.00 (s,9H, Si-tert butyl), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.62, 0.60 (2d, 3H,16-CH₃, J=7 Hz). MS-ESI (m/z): 811 (MNa⁺).

Step C. 14O-{[(1R,2R)-2-(tert-Butyl-diphenyl-silanyloxy)-4-oxo-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S) diastereomer

14-O-{[(7R,8R)-7-(tert-Butyl-diphenyl-silanyloxy)-1,4-dioxa-spiro[4.5]dec-8-ylsulfanyl]-acetyl}-mutilin+(7S,8S) diastereomer (8.03 g, 10.2 mmol) was dissolved in 100 ml ofdichloromethane and treated with montmorillonite K10(10 g) for 3 days atroom temperature. After filtration over celite the reaction mixture wasconcentrated under reduced pressure and subjected to chromatography(silica, cyclohexane/ethyl acetate 2/1) to yield 14-O-{[(1R,2R)-2-(tert-butyl-diphenyl-silanyloxy)-4-oxo-cyclohexylsulfanyl]-acetyl)-mutilin+(1S,2S) diastereomer (R_(f)=0.38, 5.24 g, 69% yield).

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 7.65-7.40 (m, 10H,aromat.-H), 6.15, 6.13 (2dd, 1H, 19-H, J=11 Hz and 18 Hz), 5.53 (d, 1H,14-H, J=7 Hz), 5.00 (m, 2H, 20-H), 4.47 (m, 1H, 11-OH), 4.24 (m, 1H,2′-H), 3.41 (t, 1H, 11-H, J=6 Hz), 3.20-3.00 (m, 2H, 22-H), 2.60 (m, 1H,1′-H), 2.40 (bs, 1H, 4-H), 1.35, 1.33 (2s, 3H, 15-CH₃), 1.06 (s, 3H,18-CH₃), 0.97 (s, 9H, Si-tert-butyl), 0.81 (d, 3H, 17-CH₃, J 7 Hz), 0.58(d, 3H, 16-CH₃, J=7 Hz).

Step D. 14-O-{[(1R,2R)-2-(tert-Butyl-diphenyl-silanyloxy)-4-hydroxyimino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S) diastereomer

To a solution of 14-O-{[(1R,2R)-2-(tert-butyl-diphenyl-silanyloxy)4-oxo-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S) diastereomer (2.50 g, 3.36 mmol) in 10 ml of dimethylformamide wasadded hydroxylamine hydrochloride (233 mg, 3.36 mmol) and triethylamine(0.47 ml, 3.36 mmol) and stirred at room temperature overnight. Thereaction mixture was concentrated under reduced pressure, diluted withwater and brine and extracted three times with ethyl acetate. Thecombined organic layers were washed twice with water and dried oversodium sulfate and filtered, The solvent was removed under reducedpressure and crude 14-O-{[(1R,2R)-2-(tert-butyl-diphenyl-silanyloxy)4-hydroxyimino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S) diastereomer (quantitative yield, cyclohexane ethyl acetate=2/1,R_(f)=0.25, 0.35) is obtained which was used for the next step withoutfurther purification.

Step E. 14-O-{[(1R, 2R,4S)-2-(tert-Butyl-diphenyl-silanyloxy)-4-hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer and 14-O-{[(1R, 2R,4R)-2-(tert-Butyl-diphenyl-silanyloxy)-4-hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4S) diastereomer

14-O-{[(1R,2R)-2-(tert-Butyl-diphenyl-silanyloxy)-4-hydroxyimino-cyclohexylsulfanyl]-acetyl},-mutilin+(1S,2S) diastereomer (2.55 g, 3.36 mmol) was dissolved in 10 ml of aceticacid and treated with sodium cyanoborohydride (210 mg, 3.36 mmol) for 90minutes at room temperature. The reaction mixture was concentrated underreduced pressure. The residue was diluted with NaHCO₃ solution andextracted three times with ethyl acetate. The combined organic layerswere dried over sodium sulfate and filtered. The filtrate was submittedto chromatography (silica, cyclohexane/ethyl acetate=2/3) to yield14-O-{[(1R, 2R,4S)-2-(tert-butyl-diphenyl-silanyloxy)-4-hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 4R) diastereomer (a) (R_(f)=0.5, 590 mg, 23% yield)and 14-O-{[(1R, 2R,4R)-2-(tert-butyl-diphenyl-silanyloxy)-4-hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4S) diastereomer (b) (R_(f)=0.3, 670 mg, 26% yield).

(a): ¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 7.6- 7.35 (m, 10H,aromat.-H), 6.93 (bs, 1H, NH/OH), 6.12, 6.08 (2dd, 1H, 19-H, J=11 Hz and18 Hz),’5.50 (m, 2H, 14-H, NH/OH), 5.00 (m, 2H, 20-H), 4.47 (m, 1H,11-OH), 3.95 (m, 1H, 2′-H), 3.40 (t, 11H, 11-H, J=6 Hz), 3.10-2.60 (m,4H, 1′-H, 4′-H, 22-H), 2.40 (bs, 1H, 4-H), 1.31, 1.30 (2s, 3H, 15-CH₃),1.00 (s, 12H, 18-CH₃, Si-tert-butyl), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.55(d, 3H, 16-CH₃, J 7 Hz).

(b): ¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 7.7-7.35 (m, 10H,aromat.-H), 6.85 (s, 1H, NH/OH), 6.16, 6.04 (2dd, 1H, 19-H, 3-11 Hz and18 Hz), 5.55 (m, 2H, 14-H, NH/OH), 5.05 (m, 2H, 20-H), 4.49 (d, 1H,11-OH, J=6 Hz), 3.55 (m, 1H, 2′-H), 3.42 (t, 1H, 11-H, J=6 Hz),AB-system (ν_(A)=3.37, ν_(B)=3.18, 22-H, J=14 Hz), 2.88 (m, 1H, 1′-H),2.54 (m, 1H, 5′-H), 2.40 (bs, 1H, 4-H), 1.39, 1.37 (2s, 3H, 15-CH₃),1.06 (s, 3H, 18-CH₃), 1.00 (s, 9H, Si-tert-butyl), 0.83 (d, 3H, 17-CH₃,J=7 Hz), 0.64, 0.62 (2d, 3H, 16-CH₃, J=7 Hz).

Step F. 14-O-{[(1R, 2R,4S)-2-(tert-Butyl-diphenyl-silanyloxy)-4-(formyl-hydroxy-amino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer

To a solution of 14-O-{[(1R, 2R,4S)-2-(tert-butyl-diphenyl-silanyloxy)-4-hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer (474 mg, 0.622 mmol) in 15 ml of tert-butyl methylether was added 2,2,2-trifluoroethyl formate (594 μl, 6.22 mmol) andheated to reflux for 5 hours. The reaction mixture was cooled to roomtemperature and added dropwise to 150 ml of heptane. The resultingprecipitate was isolated by filtration to give 14-O-{[(1R, 2R,4S)-2-(tert-butyl-diphenylsilanyloxy)-4-(formyl-hydroxy-amino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 4R) diastereomer (307 mg,62% yield, cyclohexane ethyl acetate=1/3, R_(f)=0.5) as colorless solid.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 9.6, 9.2 (2bs, 1H, NOH),8.2, 7.9 (2s, 1H, CHO), 7.65-7.35 (m, 10H, aromat.-H), 6.12, 6.08 (2dd,1H, 19-H, J=11Hz and 18 Hz), 5.50 (d, 1H, 14-H, J=7 Hz), 5.05 (m, 2H,20-H), 4.47 (m, 1H, 11-OH), 3.40 (t, 1H, 11-H, J=6 Hz), 2.37 (bs, 1H,4-H), 1.31, 1.30 (2s, 3H, 15-CH₃), 1.03 (s, 12H, 18-CH₃, Si-tert-butyl),0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.55 (d, 3H, 16-CH₃, J=6 Hz).

Step G. 14-O-{[(1R, 2R,4S)-4-(Formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer

14-O-{[(1R, 2R,4S)-2-(tert-Butyl-diphenyl-silanyloxy)-4-(formyl-hydroxy-amino-cyclohexyl-sulfanyl]-acetyl}-mutilin+(1S,2S, 4R) diastereomer (215 mg, 0.272 mmol) in 10 ml of tetrahydrofuranwas treated with tetrabutylammoniun fluoride (1.36 ml, 1M in THF, 1.36mmol) and stirred for 24 hours at room temperature. The reaction wasdiluted with a mixture of water, NaHCO3 solution and brine (1:1:1) andextracted three times with ethyl acetate. The combined organic layerswere dried over sodium sulfate, filtered and concentrated under reducedpressure. The residue was added dropwise to 250 ml heptane. Theresulting precipitate was isolated by filtration to yield 14-O-{[(1R,2R, 4S)-4-(formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1 S, 28,4R) diastereomer (97 mg, 65% yield, dichloromethane/methanol=9/1,R_(f)=0.4) as colorless solid.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 9.65, 9.25 (2bs, 1H,NOH), 8.2, 7.9 (2s, 1H, CHO), 6.13 (m, 1H, 19-H), 5.54 (d, 1H, 14-H, J=8Hz), 5.05 (m, 2H, 20-H), 4.49 (d, 1H, 11-OH, J=6 Hz), 3.42 (t, 1H, 11-H,J=6 Hz), 2.40 (bs, 1H, 4-H), 1,36 (s, 3H, 15-CH₃), 1.06 (s, 12H,18-CH₃), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 6.63 (d, 3H, 16-CH₃, J=6 Hz).MS-ESI (m/z): 574 MNa⁺), 550 (M-H)⁻, 1101 (2M-H)⁻.

Example 14 14-O-{[(1R, 2R,5S)-5-(Formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer

Step A. (7R,8R)-8-(2,4,6-Trimethylbenzylsufanyl)-1,4-dioxa-spiro[4.5]decan-7-ol+(7S,8S) diastereomer

7,8-Epoxy-1,4-dioxa-spiro[4.5]decane (Zhang, L.; Koreeda, M. OrganicLetters 2002, 4(21), 3755-3758.) (22 g, 120 mmol) was treated with2,4,6-trirethylbenzyl mercaptan (20 g, 120 mmol) according to the methodof Example 1 Step A2. After work up and chromatography of the reactionmixture (silica, cyclohexane/ethyl acetate=2/1) (7R,8R)-8-(2,4,6-trimethylbenzylsufanyl)-1,4-dioxa-spiro[4.5]decan-7-ol+(7S,8S) diastereomer (R_(f)=0.4, 33 g, 85% yield) was obtained as oil.

MS-ESI (m/z): 345 (MNa⁺), 667 (2MNa⁺).

Step B. Acetic acid (7R,8R)-7-(2,4,6-trimethylbenzylsufanyl)-1,4-dioxa-spiro[4.5]dec-8-ylester+(7S, 8S) diastereomer

To a solution of triphenylphosphine (26.5 g, 101 mmol) in 500 ml oftetrahydrofuran under argon atmosphere was added isopropylazodicarboxylate (19.6 ml, 101 mmol) and stirred for 30 minutes. Then(7R,8R)-8-(2,4,6-trimethylbenzylsufanyl)-1,4-dioxa-spiro[4.5]decan-7-ol+(7S,8S) diastereomer (27.7 g, 86 mmol) in 150 ml of tetrahydrofuran andacetic acid (7.7 ml, 135 mmol) were added and the reaction mixture washeated to 80° C. for 24 hours. The resulting reaction mixture wasconcentrated under reduced pressure and subjected to chromatography(silica, cyclohexane/ethyl acetate/methanol=3/1) to yield acetic acid(7R, 8R)-7-(2,4,6-trimethylbenzylsufanyl)-1,4-dioxa-spiro[4.5]dec-8-ylester+(7S, 8S) diastereomer (R_(f)=0.4, 7.0 g, 22% yield).

¹H NMR (500 MHz, CDCl₃, δ, ppm): 6.81 (s, 2H, aromat.-H), 4.85 (m, 1H,CHO), 3.96 (m, 4H, OCH₂CH₂O), AB-system (ν_(A)=3.83, ν_(B)=3.79, J=11Hz, SCH₂), 2.99 (m, 1H, CHS), 2.36 (s, 6H, CH₃), 2.23 (s, 3H, CH₃), 2.18(m, 1H), 2.12 (m, 1H), 2.11 (s, 3H, COCH₃), 1.90-1.58 (m, 4H). MS-ESI(m/z): 387 (MNa⁺).

Step C. (7R, 8R)-7-Mercapto-1,4-dioxa-spiro[4.5]decan-8-ol+(7S, 8S)diastereomer

Acetic acid (7R,8R)-7-(2,4,6-trimethylbenzylsufanyl)-1,4-dioxa-spiro[4.5]dec-8-ylester+(7S, 8S) diastereomer.(6.33 g, 17.4 mmol) was treated with sodium(1.6 g, 69.5 mmol) according to the method of Example 11 Step D. Afterwork up and chromatography of the reaction mixture (silica,cyclohexane/ethyl acetate=1/1) (7R,8R)-7-mercapto-1,4-dioxa-spiro[4.5]decan-8-ol+(7S, 8S) diastereomer(R_(f)=0.4, 1.36 g, 38%) was obtained.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm): 4.89 (d, 1H, OH), 3.83 (m, 4H,OCH₂CH₂O), 3.17 (m, 1H, CHO), 2.76 (m, 1H, CHS), 2.43 (s, 1H, SH),1.90-1.30, 6H). MS-ESI (m/z): 189 (M-H)⁻.

Step D. 14-O-{[(7R,8R)-8-Hydroxy-1,4-dioxa-spiro[4.5]dec-7-ylsulfanyl]-acetyl}-mutilin+{7S,8S) diastereomer

(7R, 8R)-7-Mercapto-1,4-dioxa-spiro[4.5]decan-8-ol+(7S, 8S) diastereomer(1.36 g, 7.15 mmol) was treated with pleuromutilin tosylate (3.8 g, 7.15mmol) according to the method of Example 11 Step E. After work up andchromatography of the reaction mixture (silica, cyclohexane/ethylacetate=1/1) 14-O-{[(7R,8R)-8-hydroxy-1,4-dioxa-spiro[4.5]dec-7-ylsulfanyl]-acetyl}-mutilin+(7S,8S) diastereomer (R_(f)=0.25, 1.90 g, 48%) was obtained as colorlessamorphous foam.

MS-ESI (m/z): 573 (MNa⁺), 1123 (2MNa⁺), 549 (M-H)⁻, 585 (MCl⁻).Step E. 14-O-{[(7R,8R)-8-(tert-Butyl-diphenyl-silanyloxy)-1,4-dioxa-spiro[4.5]dec-7-ylsulfanyl]-acetyl}-mutilin+(7S,8S) diastereomer

14-O-{[(7R&8R)-8-Hydroxy-1,4-dioxa-spiro[4.5]dec-7-ylsulfanyl]-acetyl}-mutilin+(7S,8S) diastereomer (1.90 g, 3.45 mmol) was treated according to the methodof Example 13 Step B. After work up and chromatography of the reactionmixture (silia, cyclohexane/ethyl acetate=3/2) 14-O-{[(7R,8R)-8-(tert-butyl-diphenyl-silanyloxy)-1,4dioxa-spiro[4. 5]dec-7-ylsulfanyl]-acetyl}-mutilin+(7S, 8S) diastereomer (R_(f)=0.6, 1.65 g,61%) was obtained as colorless amorphous foam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 7.7-7.35 (m, 10H,aromat.-H), 6.13, 6.12 (2dd, 1H, 19-H, J=11 Hz and 18 Hz), 5.53 (d, 1H,14-H, J=7 Hz), 5.05 (m, 2H, 20-H), 4.50 (m, 1H, 11-OH), 3.78 (m, 4H,OCH₂CH₂O), 3.70 (m, 1H, 2′-H), 3.42 (m, 1H, 11-H), 3.05 (m, 3H, 2′-H,22-H), 2.40 (bs, 1H, 4-H), 1.36, 1.34 (2s, 3H, 15-CH₃), 1.05 (s, 3H,18-CH₃), 1.00 (s, 9H, Si-tert-butyl), 0.81 (d, 3H, 17-CH₃, J=7 Hz),0.60, 0.58 (2d, 3H, 16-CH₃, J=7 Hz).

Step F. 14-O-{[(1R,2R)-2-(tert-Butyl-diphenyl-silanyloxy)-5-oxo-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S) diastereomer

14-O-{[(7R,8R)-8-(tert-Butyl-diphenyl-silanyloxy)-1,4-dioxa-spiro[4.5]dec-7-ylsulfanyl]-acetyl}-mutilin+(1S,8S) diastereomer (1.65 g, 2.09 mmol) was treated according to the methodof Example 13 Step C. Crude 14-O-{[(1R,2R)-2-(tert-butyl-&phenyl-silanyloxy)-5-oxo-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S)diastereomer (1.34 g, 86% yield, cyclohexane/ethyl acetate=2/1,R_(f)=0.3) was obtained as colorless amorphous foam which was directlyused for the next step.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 7.7-7.35 (m, 10H,aromat.-H), 6.11, 6.09 (2dd, 1H, 19-H, J=11 Hz and 18 Hz), 5.48 (d, 1H,14-H, J=7 Hz), 4.98 (m, 2H, 20-H), 4.47 (m, 1H, 11-OH), 4.03 (m, 1H,2′-H), 3.40 (m, 1H, 11-H), 3.04 (m, 3H, 2′-H, 22-H), 2.37 (bs, 1H, 4-H),1.31, 1.29 (2s, 3H, 15-CH₃), 1.02 (s, 12H, 18-CH₃, Si-tert-butyl), 0.81(d, 3H, 17CH₃, J=7 Hz), 0.53 (d, 3H, 16-CH₃, J=7 Hz). MS-ESI (m/z): 767(MNa⁺), 779 (MCl⁻).

Step G. 14-O-{[(1R,2R)-2-(tert-Butyl-diphenyl-silanyloxy)-5-hydroxyimino-cyclobexylsulfanyl]-acetyl}-mutilin+(1S,2S) diastereomer

14-O-{[(1R,2R)-2-(tert-Butyl-diphenyl-silanyloxy)-5-oxo-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S) diastereomer (1.34 g, 1.80 mmol) was treated according to the methodof Example 13 Step D. Crude 14-O-{[(1R,2R)-2-(tert-butyl-diphenyl-silanyloxy)-5-hydroxyimino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S) diastereomer (quantitative yield, cyclohexane/ethyl acetate=1/1,R_(f)=0.6) was obtained as colorless amorphous foam which was directlyused for the next step.

Step H. 14-O-{[(1R, 2R,5S)-2-(tert-Butyl-diphenyl-silanyloxy)-5-hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastertomer and 14-O-{[(1R, 2R,5R)-2-(Iert-Butyl-diphenyl-silanyloxy)-5-hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5S) diastereomer

14-O-{[(1R,2R)-2-(tert-butyl-diphenyl-silanyloxy)-5-hydroxyimino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S) diastereomer (2.55 g, 3.36 mmol) was treated according to the methodof Example 13 Step E. After work up and chromatography of the reactionmixture (silica, cyclohexane/ethyl acetate=1/3) 14-O-{[(1R 2R,5S)-2-(tert-butyl-diphenyl-silanyloxy)-5-hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer (a) (R_(f)=0.4, 220 mg, 16% yield) and 14-O-{[(1R,2R,5R)-2-(tert-butyl-diphenyl-silanyloxy)-5-hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5S) diastereomer (b) (R_(f)=0.25, 560 mg, 41% yield) were obtained.

(a): ¹H NMR (400 MHz, DMSO-dhd 6, δ, ppm, inter alia): 7.65-7.35 (m,19H, aromat.-H), 7.00 (bs, 1H,NH/OH), 6.11, 6.09 (2dd, 1H, 19-H, J=11 Hzand 18 Hz), 5.50 (d, 1H, 14-H, J=8 Hz), 5.00 (m, 2H, 20-H), 4.47 (m, 1H,11-OH), 3.80 (m, 1H, 2′-H), 3.40 (t 1H, 11-H, J=6 Hz), 3.00 (m, 1H,1′-H), AB-system (ν_(A)=3.93, ν_(B)=3.80, 22-H, J=15 Hz), 2.68 (m, 1H,5′-H), 2.40 (bs, 1H, 4-H), 1.31, 1.29 (2s, 3H, 15-CH₃), 1.00 (s, 12H,18-CH₃, Si-tert-butyl), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.55 (d, 3H,16-CH₃, J=7 Hz).

(b) ¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 7.7-7.35 (m, 10H,aromat.-H), 6.97 (s, 1H, NH/OH), 6.16, 6.14 (2dd, 1H, 19-H, J=11 Hz and18 Hz), 5.56 (m, 2H, 14-H, 5.40 (bs, 1H, NH/OH), 5.07 (m, 2H, 20-H),4.49, 4.48 (2d, 1H, 11-OH, J=6 Hz), 3.48 (m, 1H, 2′-H), 3.43 (t, 1H,11-H, J=6 Hz), 3.24 (m, 2H, 22-H), 2.79 (m, 1H, 1′-H), 2.40 (bs, 1H,4-H), 2.33 (m, 1H, 4′-H), 1.38, 1.35 (2s, 3H, 15-CH₃), 1.05 (s, 3H,18-CH₃), 0.98 (s, 9H, Si-tert-butyl), 0.81 (d, 3H, 17-CH₃, J=7 Hz),0.63, 0.61 (2d, 3H, 16-CH₃, J=6 Hz).

Step I. 14-O-{[(1R, 2R,5S)-2-(tert-Butyl-diphenyl-silanyloxy)-5-(formyl-hydroxy-amino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer

14-O-{[(1R, 2R,5S)-2-(tert-butyl-diphenyl-silanyloxy)-5-hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer (215 mg, 0.282 mmol) was treated according to themethod of Example 13 Step F. After isolation of the precipitate byfiltration 14-O-{[(1R, 2R,5S)-2-(tert-butyl-diphenyl-silanyloxy)-5-(formyl-hydroxy-amino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer (135 mg, 61% yield, cyclohexane/ethyl acetate 1/3,R_(f)=0.65) was obtained as colorless solid.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 9.8, 9.3 (2bs, 1H, NOH),8.2, 7.9 (2bs, 1H, CHO), 7.60-7.35 (m, 10H, aromat.-H), 6.11, 6.09 (2dd,1H, 19-H, J=11 Hz and 18 Hz), 5.50 (d, 1H, 14-H, J=8 Hz), 5.00 (m, 2H,20-H), 4.47 (d, 1H, 11-OH, J=6 Hz), 3.40 (t, 1H, 11-H, J=6 Hz), 2.37(bs, 1H, 4-H), 1.32, 1.30 (2s, 3H, 15-CH₃), 1.03 (s, 12H, 18-CH₃,Si-tert-butyl), 0.82, 0.80 (d, 3H, 17-CH₃, J=7 Hz), 0.55 (d, 3H, 16-CH₃,J=6 Hz).

Step J. 14-O-{[(1R, 2R,5S)-5-(Formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer

14-O-{[(1R, 2R,5S)-2-(tert-Butyl-diphenyl-silanyloxy)-5-(formyl-hydroxy-amino-cyclohexyl-sulfanyl]-acetyl}-mutilin+(1S, 2S, 5R) diastereomer (130 mg, 0.164 mmol) was treated according tothe method of Example 13 Step G. After isolation of the precipitate byfiltration 14-O-{[(1R, 2R,5S)-5-(formyl-hydroxy-a&rino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 5R) diastereomer (77 mg, 85% yield, dichloromethane methanol=9/1,R_(f)=0.4) was obtained as colorless solid.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 9.7, 9.3 (2bs, 1H, NOH),8.2, 7.9 (2s, iH, CHO), 6.13 (dd, 1H, 19-H, J=11 Hz and 18 Hz), 5.53 (d,1H, 14-H, J=8 Hz), 5.06 (m, 2H, 20-H), 4.91 (d, 1H, 2′-OH), 4.49 (d, 1H,1′-OH, J=6 Hz), 4.2, 3.7 (2m, 2H, 2′-H, 5′-H), 3.41 (t, 1H, 11-H, J=6Hz), 3.28 (m, 2H, 22-H), 3.13 (m, 1H, 1′-H), 2.40 (bs, 1H, 4-H), 1.35(s, 3H, 15-CH₃), 1.06 (s, 12H, 18-CH₃), 0.81 (d, 3H, 17-CH₃, J=7 Hz),0.63 (d, 3H, 16-CH₃, J=6 Hz). MS-ESI (m/z): 574 (MNa⁺), 1125 (MNa⁺), 550(M-H)-, 1101 (2M-H)⁻.

Example 15 14-O-{[(1R, 2R,3R/S)-3-(Formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3R/S) diastereomer

Step A. 14-O-{[(6R,7R)-6-Hydroxy-1,4-dioxa-spiro[4.5]dec-7-ylsulfanyl]-acetyl}-mutilin+(6S, 7S) diastereomer

6,7-Epoxy-1,4-dioxa-spiro[4.5]decane (Vankar, Y. D.; Reddy M. V.;Chaudhuri, N. C. Tetrahedron 1994, 50(37), 11057-11078.) (16.24 g, 104mmol) and pleuromutilin thiol (20.5 g, 52 mmol) were treated accordingto the method of Example 1 Step A1. After work up and chromatography ofthe reaction mixture (silica, cyclohexane/dioxane=2/1) 14-O-{[(6R,7R)-6-hydroxy-1,4-dioxa-spiro[4.5]dec-7-ylsulfanyl]-acetyl}-mutilin+(6S,7S) diastereomer (R_(f)=0.5, 15.6 g, 55% yield) was obtained ascolorless amorphous foam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.13 (dd, 1H, 19-H, J=11Hz and 18 Hz), 5.53 (d, 1H, 14-H, J=7 Hz), 5.05 (m, 2H, 20-H), 4.90 (m,1H, 2′-OH), 4.47 (m, 1H, 11-OH), 3.97 (m, 1H, 2′-H), 3.32 (m, 1H, 11-H),3.50-3.20 (m, 2H, 22-H), 2.80 (m, 1H, 1′-H), 2.40 (bs, 1H, 4-H), 1.35,1.34 (2s, 3H, 15-CH₃), 1.05 (s, 3H, 18-CH₃), 0.81 (d, 3H, 17-CH₃, J=7Hz), 0.62 (d, 3H, 16-CH₃, J=6 Hz).

Step B. 14-O-{[(1R,2R)-2-Hydroxy-3-oxo-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 25)diastereomer

14-O-{[(6R,7R)-6-Hydroxy-1,4-dioxa-spiro[4.5]dec-7-ylsulfanyl]-acetyl}-mutilin+(6S,7S) diastereomer (15.6 g, 28.4 mmol) was treated according to the methodof Example 13 Step C. After work up and chromatography of the reactionmixture (silica, cyclohexane/ethyl acetate =1/1) 14-O-{[(1R,2R)-2-hydroxy-3-oxo-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S)diastereomer (R_(f)=0.4, 3.14 g, 22% yield) was obtained as colorlessamorphous foam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 6.13 (dd, 1H, 19-H, J=11Hz and 18 Hz), 5.55 (d, 1H, 14-H, J=8 Hz), 5.23 (m, 1H, 2′-OH), 5.05 (m,2H, 20-H), 4.49 (d, 1H, 11-OH, J=6 Hz), 4.00 (m, 1H, 2′-H), 3,50-3.30(m, 3H, 11-H, 22-H), 2.86 (m, 1H, 1′-H), 2.40 (bs, 1H, 4-H), 1.35 (s,3H, 15-CH₃), 1.06 (s, 3H, 18-CH₃), 0.80 (d, 3H, 17-CH₃, J=7 Hz), 0.61(d, 3H, 16-CH₃, J=7 Hz). MS-ESI (m/z): 529 (MNa⁺), 1035 (2MNa⁺.

Step C. 14-O-([(1R,2R)-2-Hydroxy-3-hydroxyimino-cyclohexylsulfanyl]-acetyl}-mutilin +(1S,2S) diastereomer

14-O-([(1R, 2R)-2-Hydroxy-3-oxo-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S) diastereomer (3.14 g, 6.19 mmol) was treated according to the methodof Example 13 Step D. After work up and chromatography of the reactionmixture (silica, cyclohexane/ethyl acetate =1/1 ) 14-O-{[(1R,2R)-2-hydroxy-3-hydroxyimino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S) diastereomer (R_(f)=0.2, 1.75 g, 54% yield) was obtained ascolorless amorphous foam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 10.5 (s, 1H, NOH), 6.13,6.12 (2dd, 1H, 19-H,J=11 Hz and 18 Hz), 5.54 (d, 1H 14-H, J=8 Hz), 5.33(d, 1H, 2′-OH, J=4 Hz), 5.05 (m, 2H, 20-H), 4.50 (m, 1H, 11-OH), 3.96(m, 1H, 2′-H), 3.42 (t, 1H, 11-H, J=6 Hz), 3.25 (m, 2H, 22-H), 3.14 (m,1H, 1′-H), 2.40 (bs, 1H, 4-H), 1.35 (s, 3H, 15-CH₃), 1.05 (s, 3H,18-CH₃), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=7 Hz).

Step D. 14-O-{[(1R, 2R,3R/S)-2-Hydroxy-3-Hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3R/S) diastereomer

14-O-{[(1R,2R)-2-Hydroxy-3-hydroxyimino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S) diastereomer (1.75 g, 3.35 mmol) was treated according to the methodof Example 13 Step E. After work up and chromatography of the reactionmixture (silica, ethyl acetate/methanol=10/1) 14-O-{[(1R, 2R,3R_(f)=2-hydroxy-3-Hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3R/S) diastereomer (R_(f)=0.2, 1.34 g, 65% yield) was obtained ascolorless amorphous foam.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 7.1 (bs, 1H, NH/OH),6.12,6.11 (2dd, 1H, 19-H, J=11 Hz and 18 Hz), 5.53 (d, 1H, 14-H, J=8Hz), 5.05 (m, 2H, 20-H), 4.90 (m ,1H, 2′-OH), 4.5 (m, 1H, 11-OH), 3.41(t, 1H, 11-H, J=6 Hz), 3.73, 3.53, 3.30, 3.14, 3.01, 2.87 (6m, 5H, 1′-H,2′-H, 3′-H, 22-H), 2.40 (bs, 1H, 4-H), 1.35.(2s, 3H, 15-CH₃), 1.05 (s,3H, 18-CH₃), 0.81 (d, 3H, 17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=7 Hz);

Step E. 14-O-{[(1R, 2R,3R/S)-3-(Formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3R/S) diastereomer

14-O-{[(1R, 2R,3R/S)-2-hydroxy-3-Hydroxyamino-cyclohexylsulfanyl]-acetyl}-mutilin+(1S,2S, 3R/S) diastereomer (899 mg, 1.72 mmol) was treated according to themethod of Example 13 Step F. After isolation of the precipitate byfiltration 14-O- {[( 1R, 2R,3R/S)-3-(formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin+(1S, 2S, 3R/S) diastereomer (724 mg, 76% yield,dichloromethane/methanol=9/1, R_(f)=0.5) was obtained as colorlesssolid.

¹H NMR (400 MHz, DMSO-d₆, δ, ppm, inter alia): 9.6, 9.9.4, 9.1 (3bs, 1H,NOH), 8.2, 7.9 (2s, 1H, CHO), 6.13, 6.11 (2dd, 1H, 19-H, J=11Hz and 18Hz), 5.54 (d, 1H, 14-H, J=8 Hz), 5.24 (m, 1H,-2′-OH), 5.05 (m, 2H,20-H), 4.49 (m, 1H, 11-OH), 3.86, 3.60 (2m, 1H, 2′-H), 3.39 (t, 1H,11-H, J=6 Hz), 3.28, 3.13, 2.64 (3m, 4H, 1′-H, 3′-H, 22-H), 2.38 (bs,1H, 4-H), 1.36 (s, 3H, 15-CH₃), 1.06 (s, 12H, 18-CH₃), 0.81 (d, 3H,17-CH₃, J=7 Hz), 0.63 (d, 3H, 16-CH₃, J=7 Hz). MS-ESI (m/z): 574 (MNa⁺),1125 (2MNa⁺), 550 (M-H)⁻, 1101 (2M-H)⁻.

1. A compound of formula (I)

wherein n is 0 to 4; R is ethyl or vinyl; R₁ is hydrogen or (C₁₋₆)alkyl,R₂ is hydrogen or cycloalkyl including (C₃₋₆)cycloalkyl, orunsubstituted (C₁₋₆)alkyl, or (C₁₋₆)alkyl substituted by one or more ofhydroxy; preferably one or two, halogen, (C₃₋₆)cycloakyl, or R₁ ishydroxy and R₂ is formyl.
 2. A compound of formula (II)

wherein n, R₁ and R₂ are as defined in claim
 1. 3. A compound of formula(III)

wherein n, R₁ and R₂ are as defined in claim
 1. 4. A compound of formula(IV)

wherein n, R₁ and R₂ are as defined in claim
 1. 5. A compound accordingto claim 1, selected from the group consisting of 14-O-{[(1R, 2R,4R)-4-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl }-mutilin and the (1S,2S 4S) diastereomer thereof 14-O-{[(1R, 2R,5S)-5-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the (1S,2S, 5R) diastereomer thereof 14-O-{[(1R, 2R,4S)-4-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl }-mutilin and the (1S,2S, 4R) diastereomer thereof 14-O-{[(1R, 2R,5R)-5-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the (1S,2S, 5S) diastereomer thereof 14-O-{[(1R, 2R,3R)-3-Amino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the (1S,2S, 3S) diastereomer thereof 14-O-{[(1R, 2R,4R)-4-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl }-mutilin andthe (1S, 2S, 4S) diastereomer thereof 14-O-{[(1R, 2R,4R)-4-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the(1S, 2S, 4S) diastereomer thereof 14-O-{[(1R, 2R,5S)-5-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl }-mutilin and the(1S, 2S, 5R) diastereomer thereof 14-O-{[(1R, 2R,5S)-5-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl }-mutilin andthe (1S, 2S, 5R) diastereomer thereof 14-O-{[(1R, 2R,4S)-4-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl }-mutilin andthe (1S, 2S, 4R) diastereomer thereof 14-O-{[(1R, 2R,5R)-5-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl }-mutilin andthe (1S, 2S, 5S) diastereomer thereof 14-O-{[(1R, 2R,3R)-3-Ethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl }-mutilin and the(1S, 2S, 3S) diastereomer thereof 14-O-{[(1R, 2R,3R)-3-Diethylamino-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilin and the(1S, 2S, 3S) diastereomer thereof 14-O-{[(1R, 2R,4S)-4-(Formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinand the (1S, 2S, 4R) diastereomer thereof 14-O-{[(1R, 2R,5S)-5-(Formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinand the (1S, 2S, 5R) diastereomer thereof and 14-O-{[(1R, 2R,3R/S)-3-(Formyl-hydroxy-amino)-2-hydroxy-cyclohexylsulfanyl]-acetyl}-mutilinand the (1S, 2S, 3R/S) diastereomer thereof.
 6. The compound accordingto claim 1 the form of a salt and/or solvate.
 7. A compound according toclaim 1 for use as a pharmaceutical drug substance.
 8. A method oftreatment of diseases mediated by microbes which comprises administeringto a subject in need of such treatment an effective amount of a compoundof claim
 1. 9. A pharmaceutical drug composition comprising a compoundof claim 1, in association with at least one pharmaceutical excipient.10. A pharmaceutical drug composition according to claim 9, furthercomprising another pharmaceutically active agent.